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HumaClot Duoplus

| User Manual

|
Cat.No. 15650/1
Revision List of the Manual
No. DATE / Rev. REVISION DESCRIPTION
01 01/2008-03 First edition
02 02/2008-10 Adaptation for software version C5.15
03 03/2009-11 D-Dimer added
04 04/2010-07 Corrections (typographical errors, no. of figures)
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1 INTRODUCTION
This manual is considered as a part of the instrument; it has to be at the operator’s hand as well as at the
maintenance operator’s availability. For accurate installation, use and maintenance, please read the following
instructions carefully. In order to avoid instrument or personal damages, carefully read the ”GENERAL SAFETY
WARNINGS”, describing the suitable operating procedures. In case of breakdowns or any troubles with the
instrument, apply to the local Technical Service.

2 USER WARRANTY
HUMAN warrants that instruments sold by one of its authorised representatives shall be free of any defect in
material or workmanship, provided that this warranty shall apply only to defects which become apparent within
one year from the date of delivery of the new instrument to the purchaser.
The HUMAN representative shall replace or repair any defective item at no charge, except for transportation
expenses to the point of repair.
This warranty excludes the HUMAN representative from liability to replace any item considered as expendable in
the course of normal usage, e.g.: lamps, valves, syringes, glassware, fuses, diskettes, tubing etc.
The HUMAN representative shall be relieved of any liability under this warranty if the product is not used in
accordance with the manufacturer's instructions, altered in any way not specified by HUMAN, not regularly
maintained, used with equipment not approved by HUMAN or used for purposes for which it was not designed.
HUMAN shall be relieved of any obligation under this warranty, unless a completed installation / warranty
registration form is received by HUMAN within 15 days of installation of this product.
This warranty does not apply to damages incurred in shipment of goods. Any damage so incurred shall be re-ported
to the freight carrier for settlement or claim.

3 INTENDED USE OF THE INSTRUMENT [IVD]


The instrument has to be used for the expected purposes and in perfect technical conditions, by qualified
personnel, in working conditions and maintenance operations as described in this manual, according to the
GENERAL SAFETY WARNINGS. This manual contains instructions for professional qualified operators.

4 GENERAL SAFETY WARNINGS


Use only chemical reagents and accessories specified and supplied by HUMAN and/or mentioned in this manual.
Place the product so that it has proper ventilation.
The instrument should be installed on a stationary flat working surface, free from vibrations.
Do not operate in area with excessive dust.
Work at room temperature and humidity, according to the specifications listed in this manual.
Do not operate this instrument with covers and panels removed.
Only use the power cord specified for this product, with the grounding conductor of the power cord connected to
earth ground.
Use only the fuse type and rating specified by the manufacturer for this instrument, use of fuses with improper
ratings may pose electrical and fire hazards.
To avoid fire or shock hazard, observe all ratings and markings on the instrument.
Do not power the instrument in potentially explosive environment or at risk of fire.
Prior to cleaning and/or maintaining the instrument, switch off the instrument and remove the power cord.
For cleaning use only materials specified in this manual, otherwise parts may become damaged.
It is recommended always to wear protective apparel and eye protection while using this instrument.
Respective warning symbols, if appearing in this manual, should be carefully considered.

I
5 DISPOSAL MANAGEMENT CONCEPT
The currently valid local regulations governing disposal must be observed. It is in the responsibility of the user to
arrange proper disposal of the individual components.
All parts which may comprise potentially infectious materials have to be disinfected by suitable validated
procedures (autoclaving, chemical treatment) prior to disposal. Applicable local regulations for disposal have to be
carefully observed.
The Instruments and electronic accessories (without batteries, power packs etc.) must be disposed of according to
the regulations for the disposal of electronic components.
Batteries, power packs and similar power source have to be dismounted from electric/electronic parts and disposed
off in accordance with applicable local regulations.

6 INSTRUMENT DISINFECTION
Analytical instruments for in vitro diagnostic involve the handling of human samples and controls which should be
considered at least potentially infectious. Therefore every part and accessory of the respective instrument which
may have come into contact with such samples must equally be considered as potentially infectious.
Before doing any servicing on the instrument it is very important to thoroughly disinfect all possibly contaminated
parts. Before the instrument is removed from the laboratory for disposal or servicing, it must be
decontaminated/disinfected. Decontamination/disinfection should be performed by a authorised well-trained
personnel, observing all necessary safety precautions. Instruments to be returned have to be accompanied by a
disinfection certificate completed by the responsible laboratory manager. If a disinfection certificate is not
supplied, the returning laboratory will be responsible for charges resulting from non-acceptance of the instrument
by the servicing centre, or from authority’s interventions.

7 NOTICE
Every effort has been made to avoid errors in text and diagrams, however, HUMAN GmbH assumes no
responsibility for any errors which may appear in this publication. It is the policy of HUMAN GmbH to improve
products as new techniques and components become available. HUMAN GmbH therefore has to reserve the right
to change specifications if necessary in the course of such improvements.

II
NOTICE
Analytical instruments for in vitro diagnostic application involve the handling of human samples and controls
which should be considered at least potentially infectious. Therefore every part and accessory of the respective
instrument which may have come into contact with such samples must equally be considered as potentially
infectious.

BIOHAZARD

The „BIOHAZARD“ warning label must be affixed to instrument prior to first use with biological material !

Servicing Note:
Before doing any servicing on the instrument it is very important to thoroughly disinfect all possibly contaminated
parts. Before the instrument is removed from the laboratory for disposal or servicing, it must be decontaminated.
Decontamination should be performed by authorised well-trained personnel only, observing all necessary safety
precautions. Instruments to be returned have to be accompanied by a decontamination certificate completed by
the responsible laboratory manager. If a decontamination certificate is not supplied, the returning laboratory will
be responsible for charges resulting from non-acceptance of the instrument by the servicing centre, or from
authority’s interventions.

HUMAN
Gesellschaft für Biochemica und Diagnostica mbH
| Max-Planck-Ring 21 · 65205 Wiesbaden · Germany
| Tel.: +49 61 22/99 88-0 · Fax: +49 61 22/99 88-100
| e-Mail: tech-support@human.de · www.human.de

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Contents

Symbols 3
1 Safety information 3
2 General 4
2.1 Intended Purpose 4
2.2 Installation 5
2.3 Technical Data 5
3 Instrument Components 5
3.1 Incubator Block 5
3.2 Control Panel 7
3.3 Rear View of the Instrument 9
3.4 AutoHumaPette (optional) 9
3.5 Thermal-Printer (optional) 9
3.6 Barcode Scanner (optional) 9
4 THEORY OF OPERATION 10
4.1 Clotting Assay (CLOT) 11
4.2 Derived Fibrinogen (CLOT + FIB) 11
4.3 Chromogenic Assay (KINETIC) 12
4.4 Chromogenic Ecarin Assay (100 mOD) 12
4.5 Immunoturbidimetric Assay (IMMUNO) 13
5 Operating Instructions 14
5.1 Setup System 14
5.1.1 Language 14
5.1.2 Fibrinogen Concentration Units 14
5.1.3 Temperature Control 14
5.1.4 Signal 14
5.1.5 Autostart 15
5.1.6 Contrast of the LCD (Liquid Crystal Display) 15
5.1.7 Speed of the Mixer 15
5.1.8 Patient Identification 15
5.2 Setup Test 17
5.2.1 Setup Test 17
5.2.2 Units 17
5.2.3 Standard Curve 18
5.2.4 Correlation Factor (linearity index for calibration data) 18
5.2.5 Store Data 18
5.2.6 Print Test 18
5.2.7 Autostart 18
5.3 Test Analysis 20
5.3.1 Test Selection 20
5.3.2 Optic Activation & Entering Patient Identification Numbers 21
5.3.3 Duplicate testing 22
5.3.4 Starting the Analysis 22
5.3.5 Display during measuring 22
5.3.6 Manual Stop during Measurement 23
5.3.7 Return to Main Menu 23
5.3.8 Unit Key Functions 23
5.3.9 Stopwatch Functions 23
5.3.10 Result Warning Messages 23
5.4 Instrument Settings 25
5.4.1 Set System to Default 25
5.4.2 Change Temperature Adjustment 25
5.4.3 Set All Test Calibration Points to Zero 26
5.4.4 Set OD Correction 26
5.4.5 Set COAG CORRECTION 26
6 Service Menu 28
6.1 System Analysis 28
6.2 Optic Values 29
6.3 Print Sys-ID 29
7 Troubleshooting 31
8 MAINTENANCE 33
8.1 Recommended Maintenance 33
8.2 Temperature Adjustment 33
8.3 Cleaning procedures 33
9 APPLICATIONS 34
9.1 Test Overview 34
9.2 Prothrombin Time 35
9.3 Derived Fibrinogen 36
9.4 Clauss Fibrinogen Assay 36
9.5 Thrombin Time Assay 37
9.6 D-Dimer 38
9.7 APTT & APC Resistance 39
9.8 PT-Based Factor Assays (II, V, VII, X) 40
9.9 APTT-Based Factor Assays (VIII, IX, XI, XII) 41
10 SPECIAL FUNCTIONS 42
10.1 Software Upgrading 42
10.2 Interface Protocol (uni-directional) 43
11 PRODUCT CATALOGUE 43

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Symbols
Symbol Meaning Explanation

Ö Advice Indicates important information and tips.

Warning! Risk of possibly serious health hazard or damage to


equipment if warning is not heeded.

Biohazard! Danger of infection from the samples and reagents


used.

Danger! Risk to operating personnel or equipment due to electric


shock.

1 Safety information
Recommend materials
Use only original disposables.
Use only manufacturer-approved materials.
Do quality control
Carry out control measurement runs at regular intervals to ensure that the analyzer
continues to function properly.


Waste cuvettes
The cuvette blocks are intended as single-use items only.

Infectious Material
Avoid direct contact with samples and sample residues in the used cuvettes.
Infectious material such as cuvette waste and liquid waste must be disposed in compliance
with local regulations governing for infectious materials.
Wear medical infection grade protective gloves for all cleaning and maintenance work
involving potential contact with infectious liquids. Use each pair of gloves once only.
Use approved disinfectants to disinfect your hands after completion of the work.

Environmental conditions
Ambient temperature must be 18...25°C.
Humidity must be below 70%.
Avoid any vibrations or impacts to analyzer.
Do not use the analyzer in the presence of explosive or inflammable gas.

Electrical Safety
Make sure the operating voltage setting is correct before connecting the device to the power
mains.
Use only grounded electrical outlets.
Use only grounded extension cords in perfect condition. Damaged cords must be replaced
immediately.
Never interrupt protective ground contacts.
Never remove housing elements, protective covers or secured structural elements, since so
doing could expose parts carrying electric current.
Make sure surfaces such as the floor and workbench are dry while work is being done on the
device.

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2 General
The HumaClot Duoplus is a manual 2-channel photo-optical instrument that offers clotting, chromogenic &
immunoturbidimetric testing capabilities. The HumaClot Duoplus can be used for a wide variety of coagulation and
fibrinolysis tests such as:

- Prothrombin Time (PT) - D-Dimer (DD)


- Activated Prothrombin Time (aPTT) - Protein C (PC)
- Thrombin Time (TT) - Protein S (PS)
- Venom Time (VT) - von Willebrand Factor (VWF)
- Fibrinogen (FIB) - Ecarin Chromogenic Assay Thrombin (ECAT)
- Factors (FII - FXII) - Ecarin Chromogenic Assay Hirudin (ECAH)
- Antithrombin (AT3) - Plasminogen (PLG)
- Heparin (HEP) - α2-Antiplasmin (A2AP)
- Activated PC resistance (APCR)
- Lupus Anticoagulant (Screen, Confirm)

FEATURES:
- Coagulation analyzer for turbidimetric, chromogenic and immunoturbidimetric assays.
- Highly reliable, durable and nearly service-free system
- Autosensing optics to eliminate interference like bilirubin.
- Approved clotting algorithm for all kind of samples and reagents. Biphasic aPTT curve detection
- Low fibrinogen curve detection
- Fibrinogen concentration can also be derived from PT results. In addition, the standard CLAUSS method is
available.
- Calculation in % activity, INR, Ratio, g/l, ng/ml or mg/dl
- Every test is programmable with up to 5 calibration points
- Correlation analysis of calibration curves
- 2 independent stop-watch functions
- Multi-language software
- (German, English, Spanish, French, Italian, Portuguese)
- Patient identification (autoseries, manual input, barcode)
- Duplicate-test mode
- Profile testing (PT, aPTT, FIB)
- APC-R with automatic ratio calculation
- DRVVT with automatic ratio calculation
- Microvolume testing (total of 60...75 μl)
- Reagent stirring with magnetic bars
- Routines for self-checks (trouble-shooting)
- Routines for printouts (result, calibration, service, system)
- AutoHumaPette with electronically triggered start
- Automatic start triggered by reagent addition
- Optional data management and research software
- Optional printer
- Optional barcode scanner for patient identification
- Easy software update
- Interface for Laboratory Information & Management Systems (LIMS)
- Compact design and light weight

2.1 Intended Purpose


The HumaClot Duoplus is designed to carry out coagulometric tests such as PT, PTT, TT, fibrinogen, single factor tests,
chromogenic and immunoturbidimetric tests (e.g., D-dimer, antithrombin III, etc.).

Use only citrate plasma for test analysis runs: Mix 9 parts venous blood with 1 part 3.2% (0.109 M) sodium citrate
and centrifuge the mixture at 1,500 g for approx. 15 minutes. Plasma must be used within 2 hours.

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Do not use plasma with more than 25 mg/dl bilirubin concentration
Ö Do not use plasma with more than 1,000 mg/l hemoglobin concentration

The HumaClot Duoplus must be operated by a specialist trained in clinical laboratory techniques. The operator must
also have received training on this instrument and have read and understood this user manual.

2.2 Installation
No special precautions are necessary when starting up the HumaClot Duoplus. However, the following is
recommended:

- Place on a level surface in an area free from excessive temperature fluctuations.


- Avoid vibrations during measurement.
- Protect the instrument from direct sunlight, moisture and dust.
- Ensure that the mains supply conforms to the voltage and frequency rating on the instrument’s identification
plate before plugging in the instrument for the first time.

The instrument must be connected to the power supply by the mains cable supplied. If
obvious damage has occurred during shipping, do not use. Contact your local HUMAN
distributor for replacement or repair.

2.3 Technical Data


Instrument:
Boards SMD (Small Mounted Device) based
Microprocessor NEC V25
Flash-EPROM 128 KByte
RAM 128 KByte
EEPROM 2 KByte
AD-Converter 18 Bit (16 bit used)
Optics 2 LED’s ultra bright, pulse modulation control
RS 232 9600 Bd, 8 Data, 1 Stop, no parity (uni-directional)
Keyboard:
3x 8 matrixes foil keyboard, with test, function and numerical keys
with green temperature LED, indicating 37°C + 0.5°C

Display:
4 line x 20 character LCD (Liquid Crystal Display)

Incubation block:
12 cuvette pre-warming positions,
5 reagent positions
2 measuring positions

Dimensions:
WxDxH: 290x205x80 mm

Weight:
1.41 kg

Ambient temperature/humidity range for operation:


18...25°C / < 70°rH

Power Supply:
external, 42 W max
Input voltages 100 VAC to 240 VAC / 47 to 63 Hz
Output voltages +5Vdc/5A; +15Vdc/2A; -15Vdc/0.8A

3 Instrument Components
3.1 Incubator Block
The incubator block is made from aluminium, which ensures equal distribution of heat. The temperature of the
incubator block is regulated to 36.5°C - 37.5˚C

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12 positions for Measuring positions For reagent vials,
prewarming or right = Optic 2 containers or tubes:
incubation left = Optic 1 Ø 15mm
Ø 23,5mm, stirred
Ø 24,2mm

Figure 1 Incubator Block

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3.2 Control Panel

1 1

2 2

1 2 3

4 5 6

7 8 9
1 2
0 .

Figure 2 Control Panel

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Control panel in detail

ON / OFF switches the unit on and off

1 2 Optics 1 / 2 activates channel 1 and/or 2

1
Timer 1 / 2 activate timer function 1 and/or 2

Cursor up/down line up/down, select setup parameters

Menu go back to main menu or next entry

1 2 3
Numeric keys for input of calibration values and

Patient identification or
selection of submenu and
4 5 6 selection of test no.

Confirm with red enter key

7 8 9

0 .

Enter confirm entry, jump to next entry

Temperature indicates temperature is within the allowed range


of 37°C ± 0.5°C

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3.3 Rear View of the Instrument

Figure 3 Rear of Equipment

DC IN: Connection to Power Supply


Pipette: Connection to AutoHumaPette
Printer: Connection to Thermal Printer
Service: Connection to PC (Firmware upgrading, LIMS) or to barcode scanner

3.4 AutoHumaPette (optional)


Optional accessory tool for automatic test start. The pipette supports four volumes (25 to 200 μl)

3.5 Thermal-Printer (optional)


Optional accessory tool for automatic print-out. The thermal-printer must support a serial interface, set to 9600
Baud, 8 Data, 1 Stop, No parity. When the Thermal printer is connected with printer-port of the HumaClot DuoPlus,
following data will be printed automatically:

- Result Print-Out
- Test Parameter Print-Out
- Service-Report Print-Out
- System-Identification Print-Out

3.6 Barcode Scanner (optional)


Optional accessory tool for easy handling of patient identification. Up to 20 characters can be read.
Barcodes with more information will cut off at the maximum length. The barcode scanner must support a serial
interface, set to 9600 Bd, 8 Data, 1 Stop, No parity.

Ö Warning: The barcode scanner is powered with 5V over PIN 9 of the RS232
Interface of the analyzer. Do only use scanners with that feature.

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4 THEORY OF OPERATION
The HumaClot Duoplus is a highly sensitive 2-channel photometer. A very bright LED-Optic ensures accurate and
precise results, even when icteric or lipemic samples are used. The receiver signal is detected and converted to an
electrical current. During the actual test the system searches for the best signal amplification; therefore the
instrument will support a wide range of different reagents (i.e. very turbid thromboplastins or very clear reagents).
Additionally, the measurement is based on optical density (absorbance), and avoids interference by external light
effects.

CUVETTE

PLASMA + REAGENT
LASER
DETECTOR

Micro-Controller PRINTER
DISPLAY

Figure 4 The detection principle

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4.1 Clotting Assay (CLOT)
Conversion of fibrinogen to monomeric fibrin, catalysed by the enzyme thrombin, is the final reaction in the
‘coagulation cascade’. The typical clot is formed by spontaneous aggregation of monomeric fibrin molecules,
resulting in an increase in sample turbidity which is detected by the photometer. Photometric detection is started
manually by pressing the “Optic” key with simultaneous addition of the test reagent. Alternatively, the reaction is
started by the addition of the test reagent using the AutoHumaPette. The time between the start of the
photometric detection and the turning point of the reaction curve is the result. The result is displayed in seconds on
the Liquid Crystal Display (and printed automatically on the optional thermal printer.)

EXTINCTION

0.120 E END-POINT
OF REACTION

TURN-POINT
OF REACTION

Biphasic Curve

0 13.0s
START OF TEST
(i.e. PT) BEGIN OF
FIBRINOGEN-
TRANSFORMATION

Figure 5 The Turbidity Method

The diagram is representative of a typical PT curve with normal control plasma and a curve with a biphasic
reaction. Biphasic aPTT reactions are highly indicative for disseminated intravascular coagulation (DIC).

4.2 Derived Fibrinogen (CLOT + FIB)


The derived fibrinogen is determined using the clotting method described in section above. The fibrinogen
concentration in the sample is proportional to the change in optical density in the cuvette, which accompanies the
conversion of fibrinogen to fibrin at the end of the reaction. The result is expressed as “E”, which represents the
optical density at the end-point.

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4.3 Chromogenic Assay (KINETIC)
In this method, the end result is determined from the rate of optical density change. Test plasma is pre-incubated
with an enzyme (i.e. - Thrombin for determination of AT-III) and residual enzymatic activity is detected by the
addition of a chromogenic substrate. The concentration of the analyte in the test plasma is directly or indirectly
proportional (depending on the reagent system) to the rate of substrate hydrolysis, and is reported as the mean
slope of optical density per minute (delta OD(E)/min).

EXTINCTION

1.000

E2

100mOD

E1

0
t1 t3 t2 end of time in sec
test

Chromogenic Assay Chromogenic Ecarin Assay


result = slope of curve per minute result = t3
result = (E2 - E1)/(t2 - t1) if t2 - t1 = 1 min

Figure 6 The chromogenic method

4.4 Chromogenic Ecarin Assay (100 mOD)


The measurement principle is similar to standard chromogenic assays. The result, however, is expressed as the
time between the start of test and the point at which the signal reaches 100 mOD.

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4.5 Immunoturbidimetric Assay (IMMUNO)
Intense light is able to penetrate turbid solutions such as latex suspensions used for the determination of D-dimer
concentration. Latex particles designed specifically for automated D-dimer testing are coated with a monoclonal
antibody specific for D-dimer. If D-dimer antigen is present in the sample, an antigen-antibody reaction occurs,
with a simultaneously change in light transmission. The concentration of D-dimer in the sample is directly
proportional to the rate of the antigen-antibody reaction. The result is reported as the mean slope of optical
density per minute (ΔOD (E)/min, E = Extinction, a unit of light-absorbance). The following diagram illustrates the
measurement principle of D-dimer tests.

Ag - Ab OD D-DIMER
ANTIGEN
LEVEL

SAMPLE SAMPLE +
LATEX

Figure 7 Latex agglutination

The D-dimer concentration is proportional to the rate of change in optical density. The instrument calculates the
average slope of reaction, using the linear portion of the curve only.

High Dose Effect


ABSORBTION OF LIGHT

1000 ng/mL

3000 ng/mL

250 ng/mL

0
time [ s ] 100 s
maximum time

Figure 8 Relationship of light absorbance and concentration of D-dimer

The kinetic algorithm for D-dimer testing is illustrated with three typical reaction curves. At high doses the linear
relationship between signal and concentration is not valid. This is called “High Dose Hook Effect”.

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5 Operating Instructions
This section provides general instructions necessary for the user to achieve maximal use and benefit from the
HumaClot DuoPlus. Please refer to section 9.0 for specific test applications.

The On/Off switch is located on the rear panel of the instrument. For optimal results, do not operate until the
temperature indicator light is on. It takes approximately 10-15 minutes for the instrument to equilibrate to 37°C.
The general sequence of operation for test analysis is:
- Enter the “SETUP SYSTEM” submenu to confirm/change system settings
- Enter “SETUP TEST” to select test parameters and enter calibration data if desired; and
- Enter the “ANALYSIS” submenu for sample testing.

From the Main Menu, the following options are available:

1. ANALYSIS
2. SETUP TEST
3. SETUP SYSTEM
4. SERVICE

On each screen, selections are made using the Up/Down cursors. To proceed to the next menu item, press either
the “Menu” or “Enter” key. If a mistake is made, press the “Menu” key until the main menu appears and start over.

Ö To return the system to default values,


Press simultaneously “Optic 1”+ “.” + “Enter” keys.

5.1 Setup System


To enter this submenu, press #3 from the main menu. The default values for the system parameters are:

LANGUAGE: ENGLISH
FIBRINOGEN: mg/dl
TEMP.CONTROL: ON
SIGNAL: ON; VALUE 325
AUTOSTART ON
CONTRAST OF LCD: VALUE: 25
SPEED OF MIXER: VALUE: 215
PAT.IDENT.: NO PAT.ID.

5.1.1 Language
English, German, French, Italian, Spanish, Portuguese
Use the cursor keys to select, press “Enter” or “Menu” to advance.

5.1.2 Fibrinogen Concentration Units


Use the cursor keys to select mg/dl or g/l, press “Enter” or “Menu” to advance.

5.1.3 Temperature Control


On/Off - use the cursor keys to select, press “Enter“ or “Menu“ to advance. For temperature adjustment, refer to
section 8.2 Temperature Adjustment

5.1.4 Signal
ON/OFF (A beep at the start and end of testing) - use the cursor keys to select, press “Enter” or “Menu” to advance.

Higher/lower - use the cursor keys to change and “Enter” to advance.

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5.1.5 Autostart
Use the cursor keys to select and continue with “ENTER”

ON Measurement starts automatically with the addition of the reagent.


No need for pressing the optic channel key.

Off Regular mode for start with optional AutoHumaPette or manual start
by pressing relevant optic channel key.

Ö

Activate optic channel just before addition of the reagent. Movements of the cuvette can pre-start
measurement – Do not touch the cuvette if the optic channel is active!

Ö

The sensitivity of the autostart can be individually set for each test within the menu “SETUP TEST”.

5.1.6 Contrast of the LCD (Liquid Crystal Display)


Higher/lower - use the cursor keys to change and “Enter” to advance.

5.1.7 Speed of the Mixer


Higher/lower - use the cursor keys to change and “Enter” to advance.

5.1.8 Patient Identification


Four choices are available:
- No Patient ID
- Extra Input
- Autoseries
- Barcode

Use the cursor keys to select, press “Enter” or “Menu” to advance. If No Patient ID is selected, results will be printed
out without a patient identification number. If Extra Input is chosen, the user enters a patient identification
number when running each test (in the Analysis mode). The third option, Autoseries, allows the user to enter a
starting patient identification number here. Each sample run (in the Analysis mode) is then automatically
incremented by one from the starting patient identification number entered by the user. If Barcode is active, an
alphanumeric Patient ID is entered using a barcode scanner. No manual input or correction is possible. The
maximum length of the Patient ID in this mode is 20 characters. Limited by the space on the LCD, only the first 10
characters will be displayed.

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SETUP SYSTEM

1 A N A L Y S IS
2 S ETUP TEST
B IO S r e s e t 3 S ETUP SYSTEM
E E P R O M re s e t
4 S E R V IC E
MENU + EN TER O P T IC
1 + . + EN TER

LANG U AG E: ? CURSOR UP / DOW N

E N G L IS H SELEC T:
LAN GU AGE

MENU

F IB R IN O G E N IN ? C H A N G E W IT H :
CURSOR UP / DOW N

m g /d l C O N F IR M W IT H :
"E N T E R " K E Y

MENU

S E L E C T W IT H :
TEM PER ATU R E: "E N T E R " K E Y
M ARK: 3 7 .0 ° C
C H A N G E W IT H :
C U R R E N T : 3 4 .0 ° C T E M P : C U R S O R U P / D O W N + k e y "O P T IC 1 "
CONTROL: ON C ON TRO L: CU R SOR U P / D OW N

MENU

S IG N A L : C H A N G E W IT H :
CURSOR UP / DOW N

ON C O N F IR M W IT H :
"E N T E R " K E Y

MENU

AUTOSTART: C H A N G E W IT H :
CURSOR UP / DOW N

ON C O N F IR M W IT H :
"E N T E R " K E Y

MENU

CO N TRAST O F LC D : C H A N G E W IT H :
CURSOR UP / DOW N

VALU E: 25 C O N F IR M W IT H :
"E N T E R " K E Y

MENU

S P E E D O F M IX E R : C H A N G E W IT H :
CURSOR UP / DOW N

VALU E: 215 C O N F IR M W IT H :
"E N T E R " K E Y

MENU

S E L E C T W IT H :
P A T . ID E N T .: ? CURSOR UP / DOW N

IN P U T S T A R T V A L U E
A U T O S E R IE S O F A U T O S E R IE W IT H :
1000 K E Y "0 -9 "

C O N F IR M W IT H :
"E N T E R " K E Y
MENU

Figure 9 Flow diagram for the "Setup" System" Submenu

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5.2 Setup Test
The specific parameters for each test are entered in this menu. Once the initial data is entered and saved it is not
necessary to open this menu for routine testing. The submenu of “SETUP TEST” is illustrated in Figure 10. To enter
this submenu, press #2 from the main menu. The default values for “SETUP TEST” are:

METHOD: CLOTTING (or Clauss for FIB)


UNITS: seconds
CALIBRATION CURVE: Reset to zero
AUTOSTART: 800

5.2.1 Setup Test


To select a test enter the numeric code of the designated test. Alternatively, the Up/Down arrow keys can be used
to scroll through the entire test menu. For example, key in #1 to select PT. Press “Enter” to confirm selection. The
“METHOD” can also be changed when blinking.

TEST: PT

METHOD: CLOT
AUTOSTART: 800

Depending which test is active, following methods can be selected:

- CLOT Clotting assays


- CLOT+FIB Fibrinogen will be derived from PT
- CLAUSS Fibrinogen according to CLAUSS
- KINETIC Chromogenic assays
- 100mOD Chromogenic ecarin assays
- IMMUNO Immunoturbidimetric assays

5.2.2 Units
Every result is displayed in seconds [s]. However, the user can also choose to display PT results in % (% activity), R
(ratio) and I (INR). Calibration data, the mean normal PT value and/or the ISI of the thromboplastin reagent must be
entered to obtain results in %, R and I. Refer to next section for information on calibration data entry. Use the
Up/Down cursor keys to select the desired units, “Enter” to confirm and “Menu” to advance.

UNITS: PT
(s-%-R-I): s-%-I- -
NORMALVALUE: 12.2s
ISI-VALUE: 1.05

The HumaClot Duoplus reports results using the following units (which are test dependent):

E = Extinction (optical density) precision X.XXX


s = seconds precision XXX.X
R = ratio precision XX.XX
I = INR precision XX.XX
% = percent activity precision XXX.XX
U = mg/l (except FIB: mg/dl or g/l) precision XXX.X

17/44 Human HumaClot Duoplus User Manual


5.2.3 Standard Curve
To obtain results in units of concentration (ng/ml, mg/dl, IU/ml...) or % activity, a calibration curve is required with
a minimum of two and a maximum of five points. Calibration data is obtained by testing plasma [in duplicate (2) or
quadruplicate (4)] in the “Analysis” mode. An example of calibration data entry is shown below.

Example: A PT calibration curve with derived fibrinogen. Two different calibration curves are required. The order of
entry is not critical; the instrument will automatically sort calibration points.

3-point Calibration Curve for PT 4-point Calibration Curve for Derived Fib.

100% = 12.2s 591 mg/dl = 0.413E


50% = 18.0s 377 mg/dl = 0.246E
25% = 27.2s 267 mg/dl = 0.140E
0% = 0.0s 95 mg/dl = 0.042E
0% = 0.0s 0 mg/dl = 0.0E

Ö

A correct calibration curve is required to obtain results in units of concentration or % activity. Those
points remaining with no data entry are not used in the calibration curve calculation. For those tests
that require a zero calibration point a value greater than zero must be entered (i.e. 0.1% and 0.1s). All
calibration points can be reset to zero by simultaneously pressing the “0” and “Enter” keys. A full
parameter reset will eliminate calibration data for all assays.

5.2.4 Correlation Factor (linearity index for calibration data)


Insert correct calibration data and confirm storage and printing. All test-parameters, and in addition a correlation
factor, will be printed. The correlation factor (R²) indicates the linearity of the calibration curve.
It is 1.000 if the points fall into a perfectly straight line.

If R² is less than 0.980, a calibration curve with more than 3 calibration points is recommended.

5.2.5 Store Data


Yes/No. Use the cursor key to select.

5.2.6 Print Test


Yes/No. Use the cursor key to select.

5.2.7 Autostart
The sensitivity of the autostart feature can be adjusted for every test individually.
The value represents the required optical change before the instrument triggers the measurement start.

Range of sensitivity:
- Very sensitive: 300 – 500
- Normal: 500 – 1000
- Insensitive >1000

- The default value is 800.


- Increase value if test does start before adding the reagent
- Decrease value if test does not start at all.
- Setting this value to 0 will disable the autostart for the test.

Some tips:
- Pipetting clear reagent into a clear suspension will produce only low optical changes and may require a sensitive
setup ( example: FIB or chromogenic tests)
- Try to pipet directly into the suspension instead of against cuvette walls.
- Try to pipet with high pressure.

 Ö Do not use autostart below 300!


The optic could be triggered just by touching the instrument.

18/44
1 A N A L Y S IS
2 SETU P TEST
3 SETU P SYSTEM
4 S E R V IC E

MENU 2

SETU P TEST: PT

UP DOW N 0 -9 0 -9

E NTE R

TEST: PT
MENU
M ETH O D : C L O T T IN G
AU TOSTAR T: 800

MENU
C U R S O R U P /D O W N
SELEC T:
U N IT S : PT D E S IR E D U N IT S
EN TER :
(s -% -R -I): s -% -I N OR M AL VALU E
MENU
N O R M A L V A L U E : 1 2 .0 s IS I V A L U E ( o n ly fo r I)
IS I V A L U E : 1 .1 2 C O N F IR M W IT H :
"E N T E R " K E Y

MENU

O n ly d is p la ye d if % is s e le c te d !

STANDARDCURVE: PT C U R S O R U P /D O W N
1 : 9 5 .0 % = 1 2 .5 s EN TER :
2 : 3 0 .0 % = 2 7 .3 s C A L IB R A T IO N P O IN T S
C O N F IR M W IT H :
3 : 1 5 .0 % = 4 6 ,8 s "E N T E R " K E Y

MENU

PT C U R S O R U P /D O W N
STORE DATA ?
SELEC T:
YES o r N O
YE S NO

E NTE R

P R IN T T E S T : PT C U R S O R U P /D O W N

YES NO SELEC T:
YES o r N O

E NTE R

Figure 10 Flow Diagram for the "Setup Test" Submenu

19/44 Human HumaClot Duoplus User Manual


5.3 Test Analysis
To enter the submenu “Analysis”, press #1 from the main menu.

5.3.1 Test Selection


The first screen in the “Analysis” submenu will ask the user to make a test selection. This can be done two ways: by
scrolling through the test menu with the cursor Up/Down keys; or by entering a numeric test code (i.e. - “01” for PT
test). The test selection is confirmed with the “Enter” key.

TEST FOR ANALYSIS

...PT

Key Keys
UP/DOWN 0-9

enter test code


Keys
or scroll
1|4|7

Optic1:
change PT/PTT/FIB
PT: 00:00 OPTIC-1
Key
UP/DOWN scroll test PT: 00:00 OPTIC-2

Key Optic2:
3|6|9 change PT/PTT/FIB

Once a test is chosen, the system will prompt the user to remove any remaining cuvettes, and insert a filter. Once
“Enter” is pressed, the instrument performs a self-check. If any warnings or errors are identified, a message will
appear. The user is given the option to ignore the error message or warnings by pressing the “Enter” key. However,
all results will be printed with an error code and the results may be invalid. It is recommended that if an error
occurs, the test should be interrupted. Return to the Main Menu and enter the “Service” menu. Please refer to
section 6.0 for more specific instructions and information on error codes and warnings.

Ö Within the analysis menu the current test can be changed if no measurement is ongoing.

Change test optic 1 with Key “1” (PT), Key “4” (PTT), Key “7” (FIB)
Change test optic 2 with Key “3” (PT), Key “6” (PTT), Key “9” (FIB)
Change test optic 1+2 with Key “UP” , Key “DOWN”

20/44
5.3.2 Optic Activation & Entering Patient Identification Numbers
The optic can be activated in three different ways.

Single activation:
- Press optic key
- Enter / Change PID
- Press optic key again • PID is confirmed and channel is active
Repeat procedure for each desired optic channel

Multi activation with single PID:


- Press key “.” (dot)
- All channels will activated
All patient ID number will set if autoseries is active (PID = 100,101,102,103)

Multi activation with double PID:


- Press key “0”
- All channels will be activated in duplicate mode

If “Autoseries” was chosen in “Setup System”, “press the “Optic 1” key.

FIB: 100 00:00

FIB: 00:00

The patient identification number for channel one will be the number that was manually entered by the user. If
this is not the correct patient identification number, the Up/Down scroll keys can be used to increase or decrease
this number. Press the “Optic 1” key to confirm and activate the channel. Press the “Optic 2” key to continue. The
patient identification number will be automatically incremented by one for each subsequent channel.

FIB: ACTIVE 00:00

FIB: 101 00:00

If “Extra Input” was chosen in “Setup System”, the patient identification number shown for channel will be the last
entered PID. Confirm or enter a new patient ID using the numeric keyboard. Press the “Optic” key to confirm the
patient identification number and activate the channel. Repeat for the remaining channels.

To enter the patient ID with a Barcode scanner, press the "Optic 1" key ("Barcode" is displayed), scan the barcode
(the first 10 characters of the patient ID. are displayed). Continue with the next barcode.

If “No Pat ID” was selected in “Setup Test”, “No PID” will be seen for each channel. Press Optic 1 to confirm and
activate channel. Repeat for remaining channels.

21/44 Human HumaClot Duoplus User Manual


5.3.3 Duplicate testing
If duplicate testing is desired, enter the same patient identification twice. The mean result will automatically be
printed along with the individual channel data.

Ö

If the duplicates differ more than 7.5% from the mean value, the mean result will be flagged
with ‘X’!

Ö

If autoseries is selected, pressing the key “0” during optic activation will activate all channels in
duplicate mode.

5.3.4 Starting the Analysis


Place the required number of cuvettes in the cuvette pre-warming positions. To continue with the PT example,
pipette 50 μl of sample into each cuvette well. Press the “Timer 1” button to start the stopwatch. Place HEMOSTAT
THROMBOPLASTIN-SI with magnetic stirrer bar in the large centre reagent pre-warming position.

Transfer the first cuvette with sample to the measurement position. Once patient identification numbers have
been entered, press the Optic keys again to activate the channels.

FIB: ACTIVE 01:30

FIB: ACTIVE 00:26

When the timer has reached 2 minutes, pipet 100 μL of HEMOSTAT THROMBOPLASTIN-SI into the first cuvette
position. If autostart is enabled, the optic will start automatically. A beep will indicate the start of the reaction.
Repeat for the remaining cuvettes. The reaction can also be started either by pressing the optic key or by using the
AutoHumaPette, which electronically triggers the optic channel.

In summary: press the optic key once for patient identification data entry, press again to activate the channel, and
a third time to manually initiate the reaction. (Do not press a third time if using the AutoHumaPette to start the
reaction!)

 Ö If using the AutoHumaPette,


- always pipet from left to right (channel 1 - 2)!
Disable the autostart function in the SETUP SYSTEM

5.3.5 Display during measuring


Once started, a short beep is followed by a blinking screen “-------”. After the test deadtime the current optical
density will be displayed. Avoid contact with the cuvette while this message is shown. A will indicate when the
reaction is complete and the result will be displayed on the screen. If the thermal printer is attached, results will be
printed automatically.

PT: 29 mOD 01:48 PT: S 31 mOD 01:52

PT: 185 mOD 01:09 PT: R 188 mOD 01:13

Channels are in measurement.


A result has been determined on channel 2,
but channel 1 is not yet finished.
Channel 1 is running in high sensitivity

22/44
The flag “S” indicates that the analyzer has switched to high sensitivity.
The flag “R” indicates that a result was found, but it must be confirmed.

The instrument will display and sort the results. If the instrument finds a result on optic channel x, it will not
display as long as lower numbered channels are in measurement. In this case the current optical density is flagged
with “R” (result found).

5.3.6 Manual Stop during Measurement


To cancel the measurement, press both the “Enter” and “Optic” keys. This will stop the reaction. All optic channels
must be inactive in order to return to the main menu.

5.3.7 Return to Main Menu


Press key “MENU”. All optic channels must be inactive in order to return to the main menu. Stop any
measurements (refer to section above). If an optic channel is active, start and break off the measurement.

5.3.8 Unit Key Functions


Once the measurement is complete, results can be converted to units other than s, E, E/min if this option had been
selected in “Test Setup”. For each optic channel, press the corresponding “Unit” key to convert (see also section
5.2.2).

5.3.9 Stopwatch Functions


To start each stopwatch, press the “Timer” keys 1 and 2. To stop and reset, press the “Timer” key again.

5.3.10 Result Warning Messages


In addition to results, the instrument may inform the operator of unexpected sample results by attaching
status characters or error messages

DISPLAY PRINTER MEANING


* * Out of calibration (i.e. *167 %)
>|< > Out of scale (i.e. >999.9 mg/dl)
++++ NO CLOT DETECTED No clot detected within 300 seconds
---- NO CLOT DETECTED Clot detected before deadtime
???.? COAGULATION ERROR Detected reaction was not valid for coagulation
OPTIC LOW SIGNAL Light transmission was insufficient
“S” High sensitivity mode
“R” Result found
“B” Biphasic APTT found (indicative for DIC)
“F” Low fibrinogen found (indicative for liver disease)

Read section troubleshooting for further details.

23/44 Human HumaClot Duoplus User Manual


A N A L Y S IS
M A IN M E N U

1 A N A L Y S IS
2 SETU P TEST
3 SETUP SYSTEM
4 S E R V IC E

1
S E LE C T

T E S T F O R A N A L Y S IS P O S S IB L E K E Y S :
u p /d o w n ,
0 - 9,
PT

ENTER
S Y S T E M W IL L B E E X A M IN E D if a n y w a r n in g s if a n y e r r o r s
ANY
R EM O VE C U VETTE W A R N IN G E R R O R IN S Y S T E M
KEY
T E M P E R A T U R E : 3 4 .0 ° C O P T IC 1 : 0 0 0 0 0 0 0 0
O P T IC 2 : 0 0 0 0 1 0 0 0
PRESS ANY KEY PRESS ANY KEY PRESS ANY KEY

ANY
IF O K
KEY

ANY
MENU
PT : 0 0 :0 0 KEY

PT: 0 0 :0 0

O P T IC
DOT 0
1

PT: P ID 1 0 0 :0 0 PT: A C T IV E / P ID 1 0 0 :0 0 PT: A C T IV E / P ID 1 0 0 :0 0

PT: 0 0 :0 0 P T : A C T IV E / P ID 2 0 0 :0 0 PT: A C T IV E / P ID 1 0 0 :0 0

O P T IC O P T IC O P T IC
1 1 1

PT: A C T IV E / P ID 1 0 0 :0 0
O P T IC is re a d y
PT: 0 0 :0 0 to s ta rt

O P T IC OR USE
1 A U T O P IP E T T

PT: -- --- --- -- --- 0 0 :0 1 T IM E R W IL L S T A R T

PT: 0 0 :0 0
T IM E R
1

BREAK M E A S U R E M E N T IS
PT: 221 m O D 0 0 :2 2 R U N N IN G . A c tu a l o p tic a l
O P T IC d e n s ity is d is p la ye d
1 PT: 0 0 :0 0
ENTER

R E S U L T IS D IS P L A Y E D
PT: 1 2 .5 s 0 0 :0 0 A N D P R IN T E D

MENU 9 8 .5 % 1 .0 2 I T IM E R W IL L R E S E T T IM E R
PT: 0 0 :0 0 1

U N IT
1
R E S U L T S IS C O N V E R T E D
IN M E D . D IM E N S IO N S .
PT: 1 2 .5 s 0 0 :0 0 T H E D IM E N S IO N S A R E
311 U 0 .1 8 6 E S E L E C T E D IN T H E S E T U P

PT: 0 0 :0 0

Figure 11 Flow Diagram of "ANALYSIS" Submenu

24/44
5.4 Instrument Settings
5.4.1 Set System to Default
In the “main menu” press key “OPTIC 1”, “. ” (dot) and “ENTER” simultaneously.

1 ANALYSE 1 INIT TEST...


2 SETUP TEST 2 INIT BIOS ...
3 SETUP SYSTEM 3 INIT ALL ...
press simultaneously
4 SERVICE 4 TEST ACTIVATION
"OPTIC 1" + "." + "ENTER"

INIT TEST:
- All tests will be correctly entered into the memory. This means in detail that all calibration points will be reset to
zero, the method set to clotting and the units to seconds.
- The language is set to English
- The format of fibrinogen concentration is mg/dl

Ö This routine is only recommended when new software will be installed on the instrument.

INIT BIOS:
- Integration time of the receiver to 100 μs
- Temperature basis value
- LCD contrast to 33
- Speed of mixer to 215
- Frequency of beep to 325
- The serial interface is configured to 9600 Baud, Stop 1,Data 8, Parity Off

Ö This routine is only recommended when the system PCB (Main board) is replaced. The procedure must be
followed by a temperature adjustment (see section 5.4.2)!

INIT ALL:
- INIT TEST
- INIT BIOS

TEST ACTIVATION:
Allows fade in/out of tests. Use the keys “UP” ”DOWN” to change, “ENTER” to proceed and “MENU” to escape.
Deactivated test cannot be selected later in the operator’s menu.

5.4.2 Change Temperature Adjustment


When entering the position “TEMPERATURE” in the submenu “SETUP SYSTEM”, the current temperature can be
changed by pressing the keys “OPTIC 1” and “UP or DOWN” simultaneously.
The temperature will be increased or decreased in increments of 0.1°C. When pressing the keys “OPTIC 1” and “UP
or DOWN” and “1” simultaneously, the temperature will be increased or decreased in increments of 1°C. For more
information refer to section 8.2 Temperature Adjustment.

25/44 Human HumaClot Duoplus User Manual


5.4.3 Set All Test Calibration Points to Zero
When entering the position “STANDARD CURVE” in the submenu “SETUP TEST” all calibration points can be set to
zero by pressing the keys “0” and “ENTER” simultaneously.

5.4.4 Set OD Correction


Change to “TEST SETUP” and select the test which you want to correct.
When method is blinking, press key “1” and “MENU” simultaneously.

SETUP TEST: PT OD CORRECTION:

METHOD: CLOT press simultaneously +20%


"1" + "MENU"

The measured optical density of the instrument can be corrected by a factor for each test. This allows other
reagents to be adapted on the instrument.

OD-CORRECTION = 0 no effect (default)

OD-CORRECTION > 0 will cause:


- longer clotting times
- reduce sensitivity of method (more results as +++.+ s)

Positive OD-CORRECTION < 0 will cause:


- shorter clotting times
- increase sensitivity of method, which can cause wrong results (short time results!!!!)

OD-CORRECTION = -100% will invert the signal by -1.

 Ö Improper settings can cause false results. Consult your local distributor or Human GmbH before changing
the OD correction.

5.4.5 Set COAG CORRECTION


Change to “TEST SETUP” and select the test you want to correct.
When method is blinking, press key “2 and “MENU” simultaneously.

SETUP TEST: PT COAG CORRECTION:

METHOD: CLOT press simultaneously +10%


"2" + "MENU"

With the COAG CORRECTION the instrument can correct the result for better correlation to other systems or
reagents.

Example: On another instrument, plasma is measured with PT = 12.1 seconds; on the HumaClot Duoplus the result is
11.0 seconds. To get equal results, the results have to be corrected by a factor of +10%).

This can be done by entering COAG CORRECTION = 110 (Factor 1.10)


The measured optical density of the instrument can be corrected by a factor for each test. In this way it is possible
to adapt other reagents.

26/44
Negative COAG-CORRECTION will cause:
- longer clotting times
- reduce sensitivity of method (more results as +++.+ s)

Positive COAG-CORRECTION will cause:


- shorter clotting times
- increased sensitivity of method, which can cause wrong results (short time results!!!!)

 Ö A new calibration run must follow every change of COAG CORRECTION.

27/44 Human HumaClot Duoplus User Manual


6 Service Menu

1 SYSTEM ANALYSIS
2 OPTIC VALUES
3 PRINT SYS-ID
6.1 System Analysis
Press #1 from the Service submenu to enter. Perform a “SYSTEM ANALYSIS” to check instrument’s operational
status. In “SYSTEM ANALYSIS”, the HumaClot Duoplus checks the optic, temperature, memory values and analogue
to digital conversion. The error level for each channel is determined and displayed if a system error or warning is
identified.
ERROR IN SYSTEM
OPTIC 1: 00001000
OPTIC 2: 00000000
PRESS ANY KEY

An example of a “SYSTEM ANALYSIS” screen is:


The number after CH1-2 is the error code (see below) In this screen example the channel 1 LED is showing a
malfunction, while channel 2 is operating correctly.
After the self-check is complete, a service report is automatically printed.

SERVICE REPORT

HumaClot DuoPlus
N: 1000 Serial number
SOFTWARE: 1.14 Software revision
TEMP.FACTOR: 15496 Digital target
temperature value

CONTRAST: 25 LCD contract value


MIXER: 200 Speed of reagent mixer
TEMPERATURE: 37.0°C Current temperature at
reagent position

OPTIC 1. 2
SIGNAL: 32753 32679 Current optic values
(20000-35000) Allowed range of values

NOISE: 153 166 Current noise values


(0-500) Allowed range of values

SERVICE: 184 202 Current service values


(110-390) Allowed range of values

SYSTEM-ANALYSIS

OPTIC1 = 00000000 OK Errorcode


OPTIC2 = 00000000 OK Errorcode

STATISTIC Counters for different


tests

PT: 1000 1000 PTs performed on


instrument
PTT: 1000
TT: 1000
FIB: 1000
AT: 1000
Error Level
When the self-check is followed by an error message, an 8-BIT long number in binary code can be seen. Each BIT is
indicative of a specific error or warning:

28/44
temperature error ( range 36.0 - 38-0°C )
not used
not used
optic error by signal ( range: 20000 - 35000 )
optic error by service ( range: 110 - 300)
optic error by noise ( range : 0 - 500)
system error by AD conversion
system error by keyboard

Example: The Error Level “00000001” indicates that the temperature is out of range.

Refer to the troubleshooting guide for information on corrective actions.

6.2 Optic Values


From the “SERVICE” submenu, remove cuvettes and press #2 to enter “OPTIC VALUES”. The values for signal, noise
and service are displayed for each channel.

An example of the “OPTIC VALUES” screen is: CH1: 32432 (202)


152
CH2: 32169 (213)
168

Recommended values: *
SIGNAL : 31500 – 33500
NOISE: 0 - 250
SERVICE: 130 – 240

6.3 Print Sys-ID

Press #3 in the Service menu.


SYSTEM IDENTIFICATION
-----------------------------------------------------------
MAINBOARD: 955.000 MAINBOARD REVISION 955
SYSTEM: 1000 MAINBOARD INDEX 0
BIOS: 1.00 MAINBOARD SER.NO.: 1000
FLASH: 2.00 BIOS REVISION: 1
SOFTWARE: 1.14c BIOS INDEX: 0
FLASH REVISION: 2
FLASH INDEX: 0
SOFTWARE REVISION: 1
SOFTWARE INDEX: 10

29/44 Human HumaClot Duoplus User Manual


MAIN - MENU

1 ANALYSIS
2 SETUP TEST
3 SETUP SYSTEM
4 SERVICE

4 SERVICE - MENU

MENU

SERVICE
1. SYSTEM-ANALYSIS
2. OPTIC VALUES
3. PRINT SYS-ID

4 2 1

CH1: 32566 (216) REMOVE CUVETTE


113
CH2: 32635 (200)
110
PRESS ANY KEY

SYSTEM - ID ERROR-
OPTIC CHECK
WILL BE LEVELS
WILL BE SET 100 %
PRINTED

ANY
KEY SYSTEM-EXAMINATION

RESULT
OF
SYSTEM-EXAMINATION

ANY
KEY

SERVICE
SERVICE-
1. SYSTEM-ANALYSIS
REPORT WILL
2. OPTIC VALUES
ANY BE PRINTED
3. PRINT SYS-ID KEY

Figure 12 Flow Diagram of "SERVICE" Submenu

30/44
7 Troubleshooting
TEMPERATURE ERROR
DESCRIPTION Temperature is not between 36.0...38.0°C
PROBABLE CAUSE 1. Environmental temperature outside of range (draft from open window).
2. Electronic error.
3. Temperature is not adjusted correctly.
CORRECTIVE ACTION 1. Place instrument in a draft-free environment, protected from direct sun.
2. Allow instrument to warm up at least 15 min. after temp. adjustment or boot-up.
3. Adjust temperature (refer sections 8.1 & 8.2).
4. Contact your local HUMAN distributor for technical service if error persists.

OPTIC ERROR : SIGNAL


DESCRIPTION This error occurs if not enough light reaches the receiver.
The optical signal must be between 20000 – 35000.
PROBABLE CAUSE 1. Temp. out of range.
2. Dirt in optics.
3. LEDs in optic block defective.
4. Chip errors on boards.
CORRECTIVE ACTION 1. Wait until instrument has reached 37°C.
2. Clean optics (refer section 8.3).
3. Replace optic block.
4. Contact your local HUMAN distributor for technical service if error persists.

OPTIC ERROR : NOISE


DESCRIPTION Noise is produced by IC chips and external light sources (sun, lab-lighting).
The digital value of the noise must be below 500.
PROBABLE CAUSE 1. The instrument is exposed to intensive external light sources such as the sun or
halogen lamps.
2. Electronic errors.
CORRECTIVE ACTION 1. Protect instrument against sun or UV light.
2. Contact your local HUMAN distributor for technical service if error persists.

OPTIC ERROR : SERVICE


DESCRIPTION This error occurs if the required signal amplification is not between 110 – 300.
PROBABLE CAUSE 1. Temp. out of range.
2. Dirt in optics.
3. LEDs on optic block defective.
4. Chip errors on boards.
CORRECTIVE ACTION 1. Wait until instrument has reached 37°C.
2. Clean optics (refer section 8.3).
3. Replace optic block.
4. Contact your local HUMAN distributor for technical service if error persists.

OPTIC ERROR : AD-Conversion


DESCRIPTION The photo receiver converts light intensity to analogue direct current. This current is
converted to a digital signal. Every digital value has its own signature. If a value is lost, this
system error occurs.
PROBABLE CAUSE 1. CPU defective.
2. ADC defective.
CORRECTIVE ACTION Contact your local HUMAN distributor for technical service if error persists.

31/44 Human HumaClot Duoplus User Manual


ANALYSIS ERROR : ++++•
DESCRIPTION No clotting reaction is detected within 300s.
PROBABLE CAUSE 1. There really is no clot.
2. The fibrinogen concentration in the sample is below 50 mg/dl.
3. Started channel and pipetted channel are not equal.
CORRECTIVE ACTION 1. Confirm correct handling.
2. Determine clotting time with research software or observe the optical values on
the display during screen. The time until a change in signal can be observed is the
clotting time.
3. Change to HUMAN reagents.

ANALYSIS ERROR : ----


DESCRIPTION Clotting reaction starts and ends before deadtime.
PROBABLE CAUSE 1. PT based test clot before 7 sec.
2. PTT based tests clot before 15 sec.
CORRECTIVE ACTION Change to HUMAN reagents.

ANALYSIS ERROR :????


DESCRIPTION The instrument detects a reaction, but is not able to verify a clot reaction.
PROBABLE CAUSE 1. Air bubbles.
2. Touching of cuvette.
3. Clot reaction starts before deadtime.
CORRECTIVE ACTION 1. Avoid air bubbles.
2. Avoid touching the cuvette during measurement start.
3. Use of single instead of double cuvettes (reasonable for D-dimer testing)
4. Change to HUMAN reagents.

ANALYSIS ERROR : OPTIC


DESCRIPTION The received signal is below 400 digits.
PROBABLE CAUSE 1. Very turbid samples or reagents.
2. Dirt in optics.
3. Optic is defective.
CORRECTIVE ACTION 1. Clean optics.
2. Check optic value (refer section 6.2).
3. Change to HUMAN reagents.

ANALYSIS ERROR : >, <


DESCRIPTION Assay range limitation.
PROBABLE CAUSE 1. Calculated units are not within 0-999.
2. Calculated activity is not within 3 – 180%.
3. Calculated ratio is not within 0-9.
4. Calculated INR is not within 0-12.
CORRECTIVE ACTION Check test setup and recalibrate test if necessary.

ANALYSIS ERROR : *
DESCRIPTION Calculated result is extrapolated. Extrapolated results can differ from interpolated results.
PROBABLE CAUSE The obtained result is outside of calibration.
CORRECTIVE ACTION Extend the calibration curve. High and low standards are helpful.

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ANALYSIS ERROR : B
DESCRIPTION The aPTT is obtained from a biphasic reaction curve.
The “B” flag indicates DIC (no error flag)
PROBABLE CAUSE Biphasic aPTT correlates with disseminated intravascular coagulation (DIC).
CORRECTIVE ACTION Confirm DIC.
Determination of FIB, CRP and VLDL may be helpful.

ANALYSIS ERROR : F
DESCRIPTION The PT is obtained from a flat reaction curve.
The “F” – Flag is an indicator for low fibrinogen (no error flag)
PROBABLE CAUSE Fibrinogen concentration of sample is below 75 mg/dl.
Low fibrinogen levels can be caused by liver-disease or DIC.
CORRECTIVE ACTION Confirm low fibrinogen with other methods (i.e. CLAUSS method).

8 MAINTENANCE
8.1 Recommended Maintenance
Daily: Check that the optic unit and the filters are free from dirt. Clean with lint-free tissue
paper if necessary.

Monthly: Check the temperature of the incubator block. When the green LED is on, place a water filled
reagent bottle in one of the reagent positions. Insert a calibrated thermometer and record
temperature after 10 minutes. The temperature should be in the range of 36.5°C and 37.5°C.
Proceed to next section if the temperature is incorrect.

Yearly: Service check by authorized HUMAN technician (thorough cleaning of instrument, function tests,
quality control routines).

8.2 Temperature Adjustment


- When the green LED is on, fill a reagent container (bottle) with water and place in a reagent position on the
incubator block. Place a calibrated thermometer in the water.

- Allow to warm for 10 minutes.

- Enter the submenu ”Setup System“ and advance to the temperature screen. The current temperature of the
HumaClot Duo plus is displayed.

- Compare the temperature displayed by the system and the thermometer. If the temperature is different, adjust
the temperature on the HumaClot Duo plus by simultaneously pressing the Up/Down cursor keys and the “OPTIC
1” key. (To increase or decrease using larger increments, also press the numeric “1” key.)

- Wait until a stable temperature of 37.0°C is displayed on the HumaClot DuoPlus. Check and correct the system
temperature if not equivalent to the external thermometer.

- If both the thermometer and instrument display the same temperature, press the
“Enter” key and exit back to the main menu.

8.3 Cleaning procedures


The casing should be cleaned with a diluted alcohol (70%) or with a mild soap solution (no strong detergents). The
optical filter should be cleaned only with a diluted alcohol solution. Dry well with a clean, oil-free cloth.

33/44 Human HumaClot Duoplus User Manual


Warning:
- Do not use aggressive cleaning solutions!
- Do not use strong detergents material or metallic foams.

9 APPLICATIONS
9.1 Test Overview

A summary of the current test applications supported by the HumaClot Duo plus, the available medical units and
the numeric test codes are shown below:

Test Method Unit #Code


PT +/- FIB CLOT + FIB s-%-R-I-E-U 01
PTT CLOT s-R 02
TT CLOT s-R 03 GLOSSARY
FIB (CLAUSS) CLAUSS s-U 04
Method
VT CLOT s–R 05
- CLOT, clotting assay
PC CLOT + KIN s-%-U 06 - KIN, chromogenic assay
PS CLOT + KIN s-%-U 07 - IMMUNO, immunturbidimetric assay
- 100 mOD, chromogenic ecarin assay
F2 CLOT + KIN s-% 08
F5 CLOT + KIN s-% 09 Unit
- E, result in optical density
F7 CLOT + KIN s-% 10
- (Extinction); precision XXXX
F8 CLOT + KIN s-% 11 - s, result in seconds; precision XXX.X
F9 CLOT + KIN s-% 12 - %, result in activity; precision XXX.X
- U, result in mg/dl, mg/l, μg/ml, g/l;
F10 CLOT + KIN s-% 13
precision XXX (except HEP: X.XXX)
F11 CLOT + KIN s-% 14 - R, result in ratio; precision XX.XX
F12 CLOT + KIN s-% 15 - I, result in INR; precision XX.XX
F15 (Fletcher) CLOT + KIN s-% 16 #Code
HEP CLOT + KIN s–U 17 - Direct code for test selection
AT3 CLOT + KIN s-%-U 18
APCR CLOT s-%-R 19
LA-S CLOT s-%-R 26
LA-C CLOT s-%-R 27
DD IMMUNO E-U 20
VWF IMMUNO E-U 21
ECAH 100mOD E-%-U 22
ECAT 100mOD E-%-U 23
PLG KIN E-%-U 24
a2AP KIN E-%-U 25

All applications can be run with a minimum volume of 75 μl. In some cases, however, it is recommended to use
higher volumes to achieve high accuracy. The next pages suggest test procedures that have been tested on the
instrument and shown a coefficient of variation (CV) of below 5 % (depending on test, pipettes, skills of the
technician, etc.).

34/44
9.2 Prothrombin Time
REAGENT PREPARATION
Reconstitute HEMOSTAT THROMBOPLASTIN-SI according to the instructions in the package insert.

SYSTEM PREPARATION
- Turn on instrument and wait for green LED light to come on.
- Turn on printer if connected.
- Connect optional AutoHumaPette to system.
- Check setup system if necessary.
- Check setup test if necessary. Enter new calibration curve data to obtain results in % activity; enter ISI of the
Thromboplastin reagent and in-house determined mean normal PT if INR results are desired. Enter in-house
determined mean normal value if results in R are required.
- Return to main menu and enter “Analysis” by pressing #1. Select PT with the Up/Down arrow keys or enter the
numeric test code, #01. If any warning or error message appears, refer to section 7.0.

TEST PROCEDURE
Clotting Method:
- Pipette 50 μl plasma into cuvette(s).
- Pre-warm plasma for 3 min, or the time indicated in the reagent package insert. Press the “TIMER 1” key to start
stop-watch 1.
- Place HEMOSTAT THROMBOPLASTIN-SI with stirrer bar in large central reagent position.
- Transfer cuvette to measuring position.
- While incubating, press “OPTIC 1”. If selected, enter PAT-ID with numeric keys or Up/Down keys. Confirm by
pressing “OPTIC 1” again. The message “ACTIVE” is displayed and channel 1 is ready to start the reaction. Repeat
for the remaining channels.
- Add 100 μl pre-warmed HEMOSTAT THROMBOPLASTIN-SI and simultaneously press the “OPTIC 1” key. The
test will automatically start if using the AutoHumaPette. (CAUTION: When the test procedure is running,
pressing the “OPTIC 1” and the “Enter” keys will interrupt the test). Repeat for remaining channels.
- The instrument will read for 300 secs. If no clot is detected, the display will read “***”.
- The result is displayed in seconds. Press the corresponding “Unit” key for conversion of results.

ASSAY CALIBRATION
For calibration curves, a minimum of two values is required, with a maximum of 5. It is highly recommended that
more than two calibration points be used.
- Make dilutions of Normal Coagulation Reference Plasma I in 0.9% NaCl (saline). Refer to the table below for
preparation of standards.
- Determine PT with undiluted normal plasma in duplicate.
- Determine PT with diluted plasmas in duplicate.
- If no clot time is detected for the diluted plasma samples (<25%), change the sensitivity in “Setup Test” to “High”
and repeat.
- Enter calibration data, % activity and seconds.
- Check calibration curve with different controls, control levels I, II & III.

% Activity Dilution Preparation


100 none
50 1:2 200 μl plasma + 200 μl saline
25 1:4 100 μl plasma + 300 μl saline
12.5 1:8 50 μl plasma + 350 μl saline

35/44 Human HumaClot Duoplus User Manual


9.3 Derived Fibrinogen
REAGENT PREPARATION
Reconstitute HEMOSTAT THROMBOPLASTIN-SI according to the instructions in the package insert.

SYSTEM PREPARATION
- Turn on instrument and wait for green LED light to come on.
- Turn on printer if connected.
- Connect optional AutoHumaPette to system.
- Check system setup if necessary
- Check test setup if necessary. Select method clotting + fibrinogen, enter new calibration curve data.
- Return to main menu and enter “Analysis” by pressing #1. Select PT with the Up/Down arrow keys or enter the
numeric test code, #1. If any warning or error message appears, refer to section 7.0.

TEST PROCEDURE
Clotting method with fibrinogen:
- Pipette 50 μl plasma to cuvette.
- Pre-warm plasma for 2 min, or the time indicated in reagent package insert. Press the “TIMER 1” key to start stop-
watch 1.
- Place HEMOSTAT THROMBOPLASTIN-SI with stirrer bar in large central reagent position.
- Transfer cuvette to measuring position.
- While incubating, press “OPTIC 1”. If selected, enter PAT-ID with numeric keys or Up/Down keys. Confirm by
pressing “OPTIC 1” again. The message “ACTIVE” is displayed and channel 1 is ready to start the reaction. Repeat
for the remaining channels.

- Add 100 μl pre-warmed HEMOSTAT THROMBOPLASTIN-SI and simultaneously press the “OPTIC 1” key. The test
will automatically start if using the AutoHumaPette. (CAUTION: When the test procedure is running, pressing the
“OPTIC 1” and the “Enter” keys will interrupt the test). Repeat for remaining channels.
- The instrument will read for 300 secs. If no clot is detected, the display will read “***”.
- The result is displayed in seconds. Press the corresponding “Unit” key for conversion of results.

ASSAY CALIBRATION
For the derived fibrinogen test a calibration curve must be entered. It is highly recommended that more than two
calibration points be used.
- Test fibrinogen reference plasma in duplicate. For example, test fibrinogen reference plasmas with low
(∼100 mg/dl), normal (∼250 mg/dl) and high (>350 mg/dl) fibrinogen concentration. Record the absorbance
(Extinction/ OD) values for all samples.
- Enter the fibrinogen data for the calibration curve, extinction & fibrinogen values.
- Check the calibration curve with controls.
- Note: If % PT and derived fibrinogen values are required, two calibration curves must be entered.

9.4 Clauss Fibrinogen Assay


REAGENT PREPARATION
Reconstitute HEMOSTAT FIBRINOGEN (thrombin reagent and reference plasma) according to the instructions in the
kit insert.

SYSTEM PREPARATION
- Turn on instrument and wait for green LED light to come on.
- Turn on printer if connected.
- Connect optional AutoHumaPette to system.
- Check system setup if necessary.
- Check test setup if necessary and enter new calibration curve data.
- Return to main menu and enter “Analysis” by pressing #1. Select FIB with the Up/Down arrow keys or enter the
numeric test code, #6. If any warning or error message appears, refer to section 7.0.

36/44
TEST PROCEDURE
All quality control and patient samples are diluted 1:10 in Imidazole buffered saline (IBS) for testing. If the clotting
times fall outside of the linear curve, prepare and test 1:5 or 1:20 dilutions as needed.
- Pipette 100 μl of diluted sample to cuvette.
- Pre-warm sample for 4-6 min, or the time indicated in reagent package insert. Press the “TIMER 1” key to start
stop-watch 1.
- Transfer cuvette to measuring position.
- While incubating, press “OPTIC 1”. If selected, enter PAT-ID with numeric keys or Up/Down keys. Confirm by
pressing “OPTIC 1” again. The message “ACTIVE” is displayed and channel 1 is ready to start the reaction. Repeat
for the remaining channels.
- Add 50 μl Thrombin reagent (100 NIH U/ml) and simultaneously press the “OPTIC 1” key. The test will
automatically start if using the AutoHumaPette. (CAUTION: When the test procedure is running, pressing the
“OPTIC 1” and the “Enter” keys will interrupt the test).
- The instrument will read for 300 secs. If no clot is detected, the display will read “+++” and “No Clot Detected”
will print.
- The result is displayed in seconds, and both this result and the fibrinogen concentration are automatically
printed. Press the corresponding “Unit” key for fibrinogen concentration if a printer is not attached.
- For samples diluted 1:10, this is the final result. For other dilutions, the result must be corrected. For example, if
the sample was diluted 1:5, divide the result by 2; if the sample was diluted 1:20 or 1:40, multiply the result by 2
or 4, respectively.

ASSAY CALIBRATION
- Prepare standards using Normal Coagulation Reference Plasma. A suggested standard curve is shown in the
following chart:

Dilution Preparation
1:5 100 μl calibrator + 800 μl buffer
1:10 100 μl calibrator + 900 μl buffer
1:15 100 μl calibrator + 1.4 ml buffer
1:20 100 μl calibrator + 1.8 ml buffer
1:40 100 μl calibrator + 3.9 ml buffer

- Mix the dilutions and assay each in quadruplicate.


- Enter the calibration data in the “Setup Test” submenu. For example: the assigned fibrinogen value for the
plasma used to prepare the standard curve is 257 mg/dl. The 1:10 dilution corresponds to 100% activity,
therefore it is equal to 257 mg/dl. The 1:5 dilution is twice as concentrated, it is equal to 514 mg/dl. (The 1:20
dilution = 128 mg/dl and the 1: 40 = 64 mg/dl.)
- Verify the calibration curve with HEMOSTAT CONTROL PLASMA NORMAL and ABNORMAL.

9.5 Thrombin Time Assay


REAGENT PREPARATION

Reconstitute HEMOSTAT THROMBIN TIME according to the instructions in the kit insert.

SYSTEM PREPARATION
- Turn on instrument and wait for green LED light to come on.
- Turn on printer if connected.
- Connect optional AutoHumaPette to system
- Check system setup if necessary
- Check test setup if necessary. Select method clotting+fibrinogen, enter new calibration curve data.
- Return to main menu and enter “Analysis” by pressing #1. Select PT with the Up/Down arrow keys or enter the
numeric test code, #1. If any warning or error message appears, refer to section 7.0.

37/44 Human HumaClot Duoplus User Manual


TEST PROCEDURE
Clotting Method:
- Pipette 100 μl of undiluted plasma to cuvette.
- Pre-warm sample for 3 min., or the time indicated in reagent package insert. Press the “TIMER 1” key to start
stop-watch 1.
- Transfer cuvette to measuring position.
- While incubating, press “OPTIC 1”. If selected, enter PAT-ID with numeric keys or Up/Down keys. Confirm by
pressing “OPTIC 1” again. The message “ACTIVE” is displayed and channel 1 is ready to start the reaction. Repeat
for the remaining channels.
- Add 50 μl Thrombin Time Reagent and simultaneously press the “OPTIC 1” key. The test will automatically start
if using the Autopipette. (CAUTION: When the test procedure is running, pressing the “OPTIC 1” and the “Enter”
keys will interrupt the test).
The instrument will read for 300 secs. If no clot is detected, the display will read “+++” and “No Clot Detected”
will print.
- The result is displayed in seconds and is automatically printed.

9.6 D-Dimer

REAGENT PREPARATION
Refer to the kit insert for HEMOSTAT D-DIMER

SYSTEM PREPARATION
- Turn on instrument and wait for green LED light to come on.
- Turn on printer if connected.
- Check system setup if necessary.
- Check test setup if necessary and enter new calibration curve data.
- Return to main menu and enter “Analysis” by pressing #1. Select DD with the Up/Down arrow keys. If any
warning or error message appears, refer to section 7.0.

PROCEDURE
- Pipette 25 μl sample, calibrator or control into cuvette
- Add 100 μl reaction buffer
- Pre-warm for 2 – 10 minutes
- Transfer cuvette to measuring position and activate stop-watch 1
- Add 50 μl pre-warmed latex reagent
- Mix well (15 x repeated pumping by using the pipette)
- Result will be displayed ΔOD and in ng/ml
- Samples with results exceeding the reportable range (>5000 or XXX) have to be re-tested after 1 + 3 dilution with
diluents (multiply result with 4).

ASSAY CALIBRATION
Re-calibrate every 3 months.
Concentration [CAL] [DIL]
(ng/ml)
Cal. 1 3,200* 200 μl 0
Cal. 2 1,600* 200 μl 200 μl
Cal. 3 1 - -
Measure Cal. 1 and Cal. 2 in duplicate, calculate mean
and enter data into instrument.
Cal. 3 is not measured, but the zero point is entered as
1 ng/ml Æ enter 0.001 E.
* Use the lot specific value to specify the exact concentration in each dilution

Verify the calibration curve with HEMOSTAT D-DIMER CONTROL HIGH/LOW

RECOMMENDATIONS
- HumaClot Duo plus needs protection from direct sunlight (very bright external UV light will interfere with result).
- If double cuvettes are used, do not touch the cuvette during measurement. Using single cuvettes may be
reasonable.

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9.7 APTT & APC Resistance
REAGENT PREPARATION
Refer to the kit insert for HEMOSTAT aPTT-EL.

SYSTEM PREPARATION
- Turn on instrument and wait for green LED light to come on.
- Turn on printer if connected.
- Connect optional AutoHumaPette to system
- Check system setup if necessary
- Check test setup if necessary. Select method clotting + fibrinogen, enter new calibration curve data.
- Return to main menu and enter “Analysis” by pressing #1. Select PT with the Up/Down arrow keys or enter the
numeric test code, #1. If any warning or error message appears, refer to section 7.0.

TEST PROCEDURE
Clotting Method:
- Pipette 50 μl plasma to cuvette.
- Prewarm plasma for 1 - 2 min, or the time indicated in reagent package insert. Press the “TIMER 1” key to start
stop-watch 1.
- Place CaCl2 in large central reagent position.
- Add 50 μl of HemoStat aPTT-EL. Incubate for 3 - 5 minutes (refer to package insert).
- Transfer cuvette to measuring position.
- While incubating, press “OPTIC 1”. If selected, enter PAT-ID with numeric keys or Up/Down keys. Confirm by
pressing “OPTIC 1” again. The message “ACTIVE” is displayed and channel 1 is ready to start the reaction. Repeat
for the remaining channels.
- Add 50 μl pre-warmed CaCl2 and simultaneously press the “OPTIC 1” key. The test will automatically start if
using the AutoHumaPette. (CAUTION: When the test procedure is running, pressing the “OPTIC 1” and the
“Enter” keys will interrupt the test).
- The instrument will read for 300 secs. If no clot is detected, the display will read “***”.
- The result is displayed in seconds. To obtain results in R, press the “Unit 1” key.

Activated Protein C Resistance Assay (APC R):


The APCR ratio is the APTT clot time tested in the presence of APC (APTTAPC), divided by the APTT clot time in the
absence of APC:

APCR ratio = APTTAPC/APTT

The results can also be expressed as a ratio to normal control APTT, or as a % of normal control plasma.

I. APC R ratio using patient APTT value:


- Select only the unit “s” in APC test setup (#19).
- Determine the APTT of the patient plasma.
- Determine the APTT of the patient in the presence of APC using the same channel. The instrument will
automatically calculate the APC ratio. Press the corresponding Unit key for the calculated ratio. Continue from
step 2 for the next sample.

II. APCR ratio using normal control APTT value:


- Select the unit “R” in the APC test setup menu.
- Insert the APTT value of normal control plasma.
- Determine the APTT of the test sample in the presence of APC. Press the Unit key to obtain the APC ratio.

III. APC Resistance as expressed in %:

- Determine the APTT (APC) of normal control plasma. This is the 100% value for the calibration curve.
- Determine the APTT of normal control plasma, this is the 0% value.
- Select “%” in the APC Test Setup menu and insert the calibration data points.
- Determine the APTT (APC) of the test sample. Press the corresponding Unit key to obtain the result in “%”.

39/44 Human HumaClot Duoplus User Manual


9.8 PT-Based Factor Assays (II, V, VII, X)
REAGENT PREPARATION
Reconstitute Factor Deficient Plasma and Thromboplastin reagent according to the instructions in the package
insert.

SYSTEM PREPARATION
- Turn on instrument and wait for green LED light to come on.
- Turn on printer if connected.
- Connect optional AutoHumaPette to system.
- Check system setup if necessary
- Check test setup if necessary. Select method clotting+fibrinogen, enter new calibration curve data.
- Return to main menu and enter “Analysis” by pressing #1. Select PT with the Up/Down arrow keys or enter the
numeric test code, #1. If any warning or error message appears, refer to section 7.0.

TEST PROCEDURE
All quality control and patient samples are diluted 1:10 in Imidazole Buffered Saline (IBS) for testing. If the
clotting times fall outside of the linear curve, prepare and test 1:5 or 1:20 dilutions as needed. It is
recommended to test 2 dilutions (1:10 & 1:20) for each sample. Refer to the reagent package insert for more
information.

Clotting Method
- Place Thromboplastin reagent with stir bar in large central reagent position.
- Pipette 25 μl of diluted plasma and 25 μl of deficient plasma to each cuvette. Refer to chart for sample
preparation.
- Incubate for 2 min, or the time indicated in reagent package insert. Press the “TIMER 1” key to start stop-watch 1.
- Transfer cuvette to measuring position.
- While incubating, press “OPTIC 1”. If selected, enter PAT-ID with numeric keys or Up/Down keys. Confirm by
pressing “OPTIC 1” again. The message “ACTIVE” is displayed and channel 1 is ready to start the reaction. Repeat
for the remaining channels.
- Add 50 μl pre-warmed Thromboplastin reagent and simultaneously press the “OPTIC 1” key. The test will
automatically start if using the Autopipette. (CAUTION: When the test procedure is running, pressing the
“OPTIC 1” and the “Enter” keys will interrupt the test). Repeat for remaining channels.
- The instrument will read for 300 secs. If no clot is detected, the display will read “+++” and “No Clot Detected”
will print.
- The result is displayed in seconds. Press the corresponding “Unit” key for conversion of results if a printer is not
attached.
- For patient and control samples diluted 1:10, this is the final result. If other dilutions are tested, the calculated
value should be multiplied by the appropriate dilution correction factor. (i.e., samples diluted 1:20, multiply
result by 2; for 1:40 dilutions, multiply by 4, etc.)

ASSAY CALIBRATION
For calibration curves, a minimum of two values is required, with a maximum of 5. It is highly recommended that
more than two calibration points be used.

- Make dilutions of Normal Coagulation Reference Plasma in Imidazole buffered saline (IBS). A suggested standard
curve is shown below.
- Assay standards in quadruplicate as described.
- Enter calibration data (% activity and seconds) in “Setup Test”. Check calibration curve with different controls.
Sample Dilution Preparation
100% Standard 1:10 100 μl reference plasma + 900 μl IBS
50% Standard 1:20 50 μl of reference plasma + 950 μl IBS
25% Standard 1:40 25 μl of reference plasma + 975 μl IBS
12.5% Standard 1:80 500 μl of 25% standard + 500 μl IBS
Patient or Control 1:10 100 μl sample + 900 μl IBS

40/44
9.9 APTT-Based Factor Assays (VIII, IX, XI, XII)
REAGENT PREPARATION
Reconstitute Factor Deficient Plasma and APTT reagents according to package insert.

SYSTEM PREPARATION
- Turn on instrument and wait for green LED light to come on.
- Turn on printer if connected.
- Connect optional AutoHumaPette to system.
- Check system setup if necessary
- Check test setup if necessary. Select method clotting + fibrinogen, enter new calibration curve data.
- Return to main menu and enter “Analysis” by pressing #1. Select PT with the Up/Down arrow keys or enter the
numeric test code, #1. If any warning or error message appears, refer to section 7.0..

TEST PROCEDURE
All quality control and patient samples are diluted 1:10 in Imidazole Buffered Saline (IBS) for testing. If the
clotting times fall outside of the linear curve, prepare and test 1:5 or 1:20 dilutions as needed. It is
recommended to test 2 dilutions (1:10 & 1:20) for each sample. Refer to the reagent package insert for more
information.

Clotting Method
- Place CaCl2 in large central reagent position.
- Pipette 50 μl of diluted plasma and 50 uL of deficient plasma to each cuvette. Refer to chart for sample
preparation.
- Incubate for 2 min, or the time indicated in reagent package insert.
- Add 50 μl of APTT reagent. Refer to package insert for appropriate activation times. Press the “TIMER 1” key to
start stopwatch 1.
- Transfer cuvette to measuring position.
- While incubating, press “OPTIC 1”. If selected, enter PAT-ID with numeric keys or Up/Down keys. Confirm by
pressing “OPTIC 1” again. The message “ACTIVE” is displayed and channel 1 is ready to start the reaction. Repeat
for the remaining channels.
- Add 50 μl pre-warmed CaCl2 and simultaneously press the “OPTIC 1” key. The test will automatically start if
using the AutoHumaPette. (CAUTION: When the test procedure is running, pressing the “OPTIC 1” and the
“Enter” keys will interrupt the test). Repeat for remaining channels.
- The instrument will read for 300 secs. If no clot is detected, the display will read “+++” and “No Clot Detected”
will print.
- The result is displayed in seconds. Press the corresponding “Unit” key for conversion of results if a printer is not
attached.
- For patient and control samples diluted 1:10, this is the final result. If other dilutions are tested, the calculated
value should be multiplied by the appropriate dilution correction factor. (i.e., samples diluted 1:20, multiply
result by 2; for 1:40 dilutions, multiply by 4, etc.)

ASSAY CALIBRATION
For calibration curves, a minimum of two values is required, with a maximum of 5. It is highly recommended that
more than two calibration points be used.
- Make dilutions of Normal Coagulation Reference Plasma in Imidazole buffered saline (IBS). A suggested standard
curve is shown below.
- Assay standards in quadruplicate as described.
- Enter calibration data (% activity and seconds) in “Setup Test”. Check calibration curve with different controls.

Sample Dilution Preparation


100% Standard 1:10 100 μl reference plasma + 900 μl IBS
50% Standard 1:20 50 μl of reference plasma + 950 μl IBS
25% Standard 1:40 25 μl of reference plasma + 975 μl IBS
12.5% Standard 1:80 500 μl of 25% standard + 500 μl IBS
Patient or Control 1:10 100 μl sample + 900 μl IBS

41/44 Human HumaClot Duoplus User Manual


10 SPECIAL FUNCTIONS
10.1 Software Upgrading
As the process of evolution never stops, HUMAN GmbH may from time to time update the software of the
HumaClot Duo plus analyzer. The HumaClot Duo plus supports a very easy procedure for installing new revisions of
the software. For more information about latest software revision, ask your local HUMAN distributor.

Requirements:
- Windows PC,
- download cable,
- update files - flash.bat, download.exe, flashdl.exe, flashv25.exe and the analyzer firmware
xxxxx.img (i.e. cm2114.img Æ HumaClot Duo plus V1.14c)

- Save any important test setups on the instrument to paper.


- Reboot your PC. This will reset your com ports!
- Extract ”Flash Disk” to any local folder on your PC.
- Connect instrument to the PC with a null Modem cable (PIN 2-3-5 to PIN 3-2-5)
- Double click “upgrade.bat” and follow instructions.
- After a few minutes the instrument will reboot.
- Re-initialise the memory.
- Power off/on instrument.
- Press key “Optic 1” +”.” +”ENTER” simultaneously -> message “INIT SYSTEM…” should appear and afterwards
instrument should reboot automatically.
- Restore test setup parameters manually.

Problems during updates:


Update hangs at the following screen:

Æ Reboot PC and try again.


Æ If transfer still fails, boot PC with MS DOS floppy disk and perform MS DOS update.

- Nothing happens after clicking “upgrade.bat”.


Æ Perform update with MS DOS commands.

- MS DOS update (for Windows 2000 / XP)

- Open Windows Explorer and extract all files to, for example, drive “C”, folder “HUMAN”.
- Go to MS Windows “START/RUN” and enter “cmd” -> DOS command window will open
- Change to directory with command “cd c:\human”
- Enter command “dir” -> all files including upgrade.bat should be listed
- Enter command “upgrade.bat” -> a dialog window should open.
- Follow instructions. Update will start and when finished the instrument will reboot.

42/44
10.2 Interface Protocol (uni-directional)
Protocol: TECAM = Monitoring
Instrument: HumaClot Duo Plus
Interface: 9600 Bd, no parity, 8 bit. 1 stop bit
Transmission: result data = yes
Reaction curve data = yes

ASCII command characters:


STX start of transmission asc(2)
ETX end of transmission asc(3)
TAB vertical tabulator asc(9)
LF line feed asc(10)
CR carriage return asc(13)
DLE idle asc(16)

Overview of records and principle:

Curve Request Record

Curve Start Record

HumaClot
Coatron
Duo plus Result Record PC

Curve Record

Curve Undo Record

Figure 3 LIMS communication

The result record is always transmitted from the HumaClot DuoPlus.


The curve record transmission must be activated from PC with a request record.

Æ For more details, please read separate description

11 PRODUCT CATALOGUE
HumaClot Duo plus (with Standard Package)
REF: 15650

Including: 1 pc HumaClot DuoPlus


1 pc Power Supply 100-240 Vac
1 pc Power cord (Europe)
1 pc Reagent adapter Øa 23,4 - Øi 16,0 mm
1 pc Reagent adapter Øa 24,1 - Øi 22,6 mm
1 pc Printer cable
1 pc Stirring magnet
5 pc Reagent container
25 pcs Double cuvettes
1 pc User Manual
1 pc Warranty card
1 pc Label ”Biohazard“
1 pc Sheet ”Biohazard“

43/44 Human HumaClot Duoplus User Manual


Consumables, Accessories and Reagents
REF Description Content Qty

15651/10 Double-cuvette (2 pos/ea) 250 1 Pack


18690 Single cuvettes 500 1 Pack
15651/11 Reagent adapter Øa 23.4 - Øi 16.0 mm 1 1 Pc
15651/12 Reagent adapter Øa 24.1 - Øi 22.6 mm 1 1 Pc
15651/13 Stirrer bars 4 1 Pack
15651/14 Power cord Europe 1 1 Pc
15651/15 Power cord US 1 1 Pc
15651/16 Reagent container • 22.5 mm 100 1 Pack
15651/17 Reagent tubes • 16 mm 100 1 Pac
15651/20 Thermal printer 230 Vac 1 1 Pc
15651/21 Thermal printer 120 Vac 1 1 Pc
19910/20 Pipette tips 25/50/100/200 μl 1.000 1 Pack
15651/25 Thermal paper, 57mm, for printer 15651/20 5 1 Pack
15651/26 Printer cable for printer, long 1 1 Pc
15651/60 CCD- Barcode scanner 1 1 Pc

19515A AutoHumaPette 25, 50, 100 and 200 μl 1 Pc

HEMOSTAT THROMBOPLASTIN-SI
Thromboplastin for the determination of Prothrombin Time (Quick test, PT)

Cat. No. Format Unit / Size


31002 Complete kit 6 x 2 ml
31003 Complete kit 6 x 10 ml

HEMOSTAT aPTT-EL
Activated Partial Thromboplastin Time

Cat. No. Format Unit / Size


33002 Complete kit 2 x 6 x 4 ml
33012 aPTT reagent 6 x 4 ml
33013 aPTT reagent 6 x 10 ml
33022 CaCl2 4 x 30 ml

HEMOSTAT FIBRINOGEN
Thrombin reagent for the determination of Fibrinogen

Cat. No. Format Unit / Size


33002 Complete kit 5 x 2 ml

HEMOSTAT THROMBIN TIME


Thrombin reagent for the manual and automated determination of the Thrombin Time

Cat. No. Format Unit / Size


34002 Complete kit 6 x 1 ml

HEMOSTAT CONTROL PLASMA


Normal and abnormal control plasmas with target values for PT, PTT, FIB, TT

Cat. No. Format Unit / Size


35001 CONTROL PLASMA NORMAL 6 x 1 ml
35002 CONTROL PLASMA ABNORMAL 6 x 1 ml

HEMOSTAT D-DIMER
Latex reagent, reaction buffer, sample diluent and calibrator for determination of D-dimer

Cat. No. Format Unit / Size


36002 Complete kit 2 x 20 tests

HEMOSTAT D-DIMER CONTROL HIGH/LOW


Control plasma with high and low D-dimer concentration

Cat. No. Format Unit / Size


36012 2 x high, 2 x low 4 x 1 ml

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HUMAN
Gesellschaft für Biochemica und Diagnostica mbH
| Max-Planck-Ring 21 · 65205 Wiesbaden · Germany
| Tel.: +49 61 22/99 88-0 · Fax: +49 61 22/99 88-100
| e-Mail: human@human.de · www.human.de