Methods of Counting Cells

• Manual

• Impedance

• Optical
 Manual Counting Method
WBC/RBC/Platelets
• Dilutions of Blood were made and Cells Counted

in a Counting Chamber.

Differential
• A Blood Smear is Stained and the Different

Types of Cell Identified & Counted.

 Impedance Counting Method
A Regulated Current Between Two
Electrodes in a Diluent

 The Electrodes are Insulated Except

for a Small Aperture

Cells Passing through the Aperture

Break the Circuit

Signals are Interpreted as Number and

Size
 Optical Counting Method
• Cells Suspended in a Diluent are
Pumped Through a Glass Cell onto
which a Light Beam is Focused

• Cells Intercept the Beam of Light

(Light is Scattered and Absorbed)

• Signals are Interpreted as

Number, Size and Density
SHUTTLE SHEATH STREAM
CHAMBER
SAMPLE STREAM
 Complete Blood Count CBC /
• WBC FBC
Total White Cell Count
• RBC Total Red Blood Cell Count
• HGB Haemoglobin
• HCT Haematocrit / Packed Cell Volume
• MCV Mean Cell Volume
• MCH Mean Cell Hgb
• MCHC Mean Cell Hgb Concentration
• CHCM Cell Hgb Concentration Mean
• HDW Haemoglobin Distribution Width
• RDW Red Cell Distribution Width
• PLT Total Platelet Count
 1. Autosampler
2. Manual Open-tube Sampler
3. Manual Closed-tube Sampler
5
6

4
8 4. Unified Fluidics Circuit
7 5. Perox Optical Assembly
2 3
6. Rbc Optical Assembly
9 7. Reagents, Sheath, Wash &
1 Rinse
8. Vacuum & Pressure Regulator
Knobs
9. Touchpad
Barcode Reader
Waste Removal System
The Touchpad
Unified Fluidics Circuit
UFC
UFC

1. Hgb Colorimeter

2. Baso Reaction Chamber

3. RBC Reaction Chamber

4. Retics Reaction Chamber

5. Perox Reaction Chamber

6. Sample Shear Valve

7. Reagent Pump Assembly

Sample Shear Valve

As V72 opens sample

enters shear valve
at point 1.
CBC aliquots taken.

If V73 used instead then

different aliquot in perox
channel is also taken.

V74 adds retic aliquot

 Stand- Alone Automatic
Waste Container Waste Removal

1 2
3

4
2
3

5
Reagent Inventory
Advia 120 Methods

The Haemoglobin Method

 The Haemoglobin Method

HGB reaction chamber

Lamp
assembly UFC Filter + Photodiode

Optical Readings are Obtained Colorimetrically at 546nm

 The Haemoglobin Bed
1. Advia 120 Sheath/rinse Reading from Previous Cycle
% Transmission

2. Draining and Refilling with the Reaction Solution

3. Reaction Solution Readings (Sample Mean)

4. Draining and Refilling with the Advia 120 Sheath rinse

Seconds
5. Advia 120 Sheath rinse Readings (Baseline Mean)
Calculating Reported Parameters
• HGB Log (Sample Mean) x Hgb Cal
Factor x 50.0 (Haemoglobin) (Baseline Mean)
• MCH (Mean Corpuscular Haemoglobin)
(HGB) x 10
(RBC)
• MCHC (Mean Corpuscular Haemoglobin
Concentration)
(HGB ) x 1000 ([ RBC x MCV])
Advia 120 Methods

The Perox Method

 The Perox Method
Absorption
detector

Filter
Sample Scatter
detector
Tungsten Stream
lamp

Dark
Beam
splitter stop
The Perox Method
Scatter signal to measure the
volume of the cells

Absorption signal for Peroxidase

activity measurement

Cells with medium Peroxidase

activity absorbs less light

than cells with high Peroxidase

activity
 Peroxidase Cytogram
Y 1 Noise

7 2 nRBC

5 6 3 Platelet Clumps

4
Cell Size

Lymphocyte & Basophil

5 Luc

4 6 Monocytes

2 3 7 Neutrophils
2 8
1 8 Eosinophils

Peroxidase Activity X
Advia 120 Methods

The RBC Method

The RBC Method

No matter
what your shape
or size ....
We can make you
a SPHERE
 RBC’s Biconcave Disc

RBC’s Sphered
Isovolumetrically Sphered RBC’s
and Plts
 Orientation of Cells is
Not a Variable
 Predictable Light Scatter and
Absorption
 The Flow Cell
Advia 120 Sheath/rinse Encases the

Sample Stream as the Two Fluids Pass

Through the Flowcell

Cells are in Laminar flow, they file past

the Light source in single file

Shuttle Chamber S heath Stream

Sample
Sample stream Concentric Flow Cell Module (CFM)
 The RBC Method

Laser Light

Low Angle Scatter 2o - 3o (Volume)

Laser Light

High Angle Scatter 5o - 15o (Hgb Concentration)

 Laser Bench
Sample Dark
Laser diode Beam-splitter Mirror
Stream Stop

Absorption Low-angle High-angle

Detector Detector Detector

Front view of dark stop

 RBC/PLT Scatter
High Angle Detector
Measures RBC
Density/Hgb content

Laser
Zero Angle

Low Angle Detector

Measures size
 RBC Volume/Haemoglobin
Concentration Cytogram
Volume / Haemoglobin Concentration
(V/HC) Cytogram is a Linear Version
of the RBC Mie Map that Appears on
Volume

the RBC Cytogram. (Tic-Tac-Toe)

1. 60 fL Volume marker

2. 120 fL Volume marker

3. 28 g/dL HC marker

4. 41 g/dL HC marker
Haemoglobin Concentration