KTA system Training Guide

Purifier 100 / 10

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KTApurifier

Buffers
Injector

Box
Monitor

On-line
Filter

Conductivity
pH

Outlet Valve

UV Detector

Mixer

Pumps

Pump A
Pump B

Buffer switching
valve

KTApurifier 10 / 100
In-line
filter

1 7
6
2
3 4 5

Injection
valve (V1)

UV

3 Wavelength
detector

Cond.

Conductivity
monitor

pH

Mixer

pH detector

System
pumps

Fraction
collector
Frac-950

2
3

7
6
5

Outlet
valve (V4)

KTApurifier + Sample Pump + Sample Valve

In-line
filter
Injection
valve (V1)
1 7
6
2
3 45

Mixer

UV

3 Wavelength
detector

Cond

Conductivity
monitor

System
pumps

pH

Sample
pump

pH detector

Fraction
collector
Frac-950

1 87
2
6
345

1 87
2
6
345

Sample selection
valve (V5)

Outlet
valve (V4)

KTApurifier 100 Air

In-line
filter

1 7
6
2
3 4 5

Injection
valve (V1)

UV

3 Wavelength
detector

Cond.

Conductivity
monitor

pH

Mixer

pH detector

System
pumps

Fraction
collector
Frac-950

Air
sensor

1
2
3

7
6
5

Outlet
valve (V4)

KTApurifier + Buffer Valve

In-line
filter

1 7
6
2
3 4 5

Injection
valve (V1)

UV

3 Wavelength
detector

Cond.

Conductivity
monitor

pH

Mixer

pH detector

System
pumps

Fraction
collector
Frac-950

2
3

1 87
2
6
345

Buffer
select
valve (V6)

7
6
5

Outlet
valve (V4)

KTApurifier 10 XT
In-line
filter

1 7
6
2
3 4 5

Injection
valve (V1)

UV

3 Wavelength
detector

Cond.

Conductivity
monitor

pH

Mixer

pH detector

System
pumps

Fraction
collector
Frac-950

2
3

7
6
5

Outlet
valve (V4)

PUMP P-900

Mixer
Pump A

Pump B

Proportion
Valve
A2

Acid /Alkali

KTA 10 mls/min
KTA 100 mls/min

A1 B2

Buffer

B1

Salt

GRADIENT
<6 mls/min
6 -14 mls/min
<30 mls/min
30 - 50 mls/min
50 - 100 mls/min

Water

MIXER
0.6 ml mixer
2 ml mixer
2 mls mixer
5 mls mixer
12 mls mixer

PUMP P-900 Inlets

Mixer

Pump B

Pump A

Switching
Valves
Buffer Select
Valve IV-908

A2

A1

2
3

A11

A12

A13

1 8

A14

B1

Water

7
5

A16
A15

B2

A17

A18

Salt

PUMP P-900 Manually Priming Buffer Inlet Lines

Pump A

Common inlet

A2

Pump B

A1

Common inlet

Manual

Flowpath

Manual

A1 or A2

BufferValveA B1 or B2

B1

B2

Flowpath
BufferValveB

Open

Draw A
ir
out of
Buffer
Line

PUMP P-900 Back - Wash Lines

Pump Back-Wash
Lines

Syringe fitting
Wash OUT
IN-LINE
one-way
valve

Buffer
OUT
Ball outlet
check valve

High pressure
Piston seal
Piston

20% Ethanol : Water

Backwash pump
diaphragm

Ball inlet
check valve

Wash
IN

Buffer
IN

ON-LINE FILTERS
P-903 ( 10 ml/min Pump )
Low Flow HPLC Filter

P-901 ( 100 ml/min Pump )

High Flow Fibre Filter
Polymer Support

LOAD (sample)

INJECT (sample)

2
3

Pump

6
5

Waste

3
4

Sample

8
7

PV-908
1

8
7

7
2

Pump

7
3

PV-908
1

Sample

Pump

7
3

6
5

PV-908

Column

Column

Column

LOAD (wash loop)

Waste

Sample

Waste

INJECTION VALVE INV- 907

1

COLUMN

2
LOAD
SAMPLE

PUMP

Waste

Waste

LOAD
1

COLUMN

PUMP

3
4

6
5

Waste

Waste

INJECT
1

COLUMN

PUMP
3
4

Waste

6
5

Waste

WASTE

SAMPLE LOADING METHOD

1

COLUMN
2

LOAD
SAMPLE

PUMP

3
6
5

4
Needle Port

Waste

Waste

Always tighten by hand

with needle inserted

Do Not Overtighten

COLUMN
LOAD
SAMPLE

PUMP

3
6
4

Luer Lock
Adapter

Waste

Waste

Superloop

SUPERLOOPS

10 ml
50 ml
Buffer

150 ml

Plunger

BUFFER
FLUSH

SAMPLE
LOAD

SAMPLE
INJECT

SAMPLE
FLUSH

WASTE

PUMP or
SYRINGE

COLUMN

COLUMN

Sample
Etched
Groove

Volume

Max. allowed column

pressure

Code No.

10 ml
50 ml
150 ml

4 Mpa
4 Mpa
2 Mpa

18-1113-81
18-1113-82
18-1023-85

INV-907 Injection Valve - Injection Port

PEEK Valve Body
Injection Needle Port

Valve Seal
Ferrule

NeedleTube

Needle

Needle
Adapter

INV-907 Injection Valve - Luer Adapter

PEEK
PEEKInjection
InjectionValve
ValveBody
Body
PEEK
PEEKInjection
InjectionLuer
LuerAdaptor
Adaptor

Valve
ValveSeal
Seal

Valve
ValveChannel
ChannelPlate
Plate

Fluid Dynamics

Flow
PEEK Tubing
High Resistance
to Flow

Low Resistance
to Flow

End

Beginning

Fluid Dynamics

Flow - Injecting a Loop

The way we imagine it Flows

The way it Actually Flows

Bubble Segmentation

100ul Loop
Waste

50ul Load

60ul Load
100ul Load

200ul Load

Partial Loading Loop = No more than 1/2 Loop Volume

Total Loading Loop = Inject 2X Loop Volume

LOADING A SAMPLE LOOP

Injection valve
INV-907

Column

Load

Inject

Sample

Sample
pump
P-950

Pump

6
5

Waste

LOADING A SUPER LOOP

Injection valve
INV-907

Column

Load
Inject

Pump
6

Buffer

Sample
Waste

Sample

MULTI-LOADING A SAMPLE LOOP

Injection valve
INV-907

Column

Load

Inject

Pump
6

PV-908
1

2
3

I/O
4

7
6

Wash
Samples

Waste

MULTI-LOADING A SUPER LOOP

Column

1
2

Pump

Load
Inject

Injection valve
INV-907
7
3

Waste

6
5

PV-908
8
1
2
3

I/O
4

7
6

Samples
Wash
S3 S4 S5 S6

S8

DIRECT LOADING ON TO COLUMN

Column

Pump

2
3

Inject
Load

6
4

Extra Tube

1/16 or 1/8
Tubing

Waste
Sample

REVERSE FLOW VALVE ( INV-907 )

1
7

Pump

Column

Extra Tube

Stop
Plug

MONITOR

Downward Flow
Position 1
1
7

Pump

3
4

Column

Extra Tube

Stop
Plug

MONITOR

Upward Flow
Position 2

OUTLET VALVE PV-908

DETECTOR
To Fraction
Collector
Default

F8

F1
F2
F3

I/O
F4

F5

F7
F6

F7
Waste

F8

F4
F3

F5

F6

Fraction Collection
Flowthrough
Fractionation

Peak
Fractionation

Eluate
Fractionation

100%B

0%B

mAU

100

50

Waste

End

D
N
U
O
B
N
U

D
N
U
O
B
Start

Inject

500

250

%B

A1 A2 A3.

HiTrap Connections
1/16 Female

1/16 Male
18-1112-55

1/16 Fingertight Fitting

1/16 Female

M6 Male
18-1112-57

M6 male / 1/16 Female

Phenyl Sepharose

HiTrap Column
M6 Male
M6 Female
18-1112-58

M6 female / 1/16 male

Flow Cell Inlet

1/16 Male

Multiple HiTrap Arrangment ( Series )

Phenyl Sepharose

Phenyl Sepharose

Phenyl Sepharose

Phenyl Sepharose

Phenyl Sepharose

Phenyl Sepharose

De

or
t
c
te

t or
c
e
t
De

or
t
c
te
De

Traditional FPLC fitting to KTA / HPLC

Injector

1/16 Fingertight Fitting

18-1112-55

10
0u

1/16 FEMALE / M6 FEMALE

18-1123-94

Flange M6 Fitting

1/16 MALE / M6 FEMALE

18-1112-58

Flow Cell

Tubing Fittings
18-1112-55

1/16 Fingertight Fitting

1/16 PEEK

TM

Tubing

18-1112-58

Flange Fitting Adapter

FPLC Flange Fitting

18-1112-51

Luer Fitting Adapter

18-1112-52

Fingertight Stop Plug

Flangless Inlet Connector

3/16
18-1112-49
1/8
18-1121-17
1/16
18-1127-07

18-1112-57

M6 male / 1/16 Female

Luer Syringe

18-1112-50
5/16 Stop Plug

Flangless Buffer Inlet

Ferrule
3/16
18-1112-48
1/8
18-1121-18
1/16
18-1127-06

18-1017-98
Tubing Connector
M6 male / O - ring

18-3955-01

1/16female X 1/16female Union Ti

Union + 2 Fingertight Fittings

18-1123-94

M6 female X 1/16female Union

FPLC Flange - 1/16 Fingertight

COMPRESSION FITTINGS

Untightened

Correctly tightened
Obstructed

Over tightened Tubing Restricted

PEEK Tubing
TM

Gray, 1/16x 1.0 mm ( 7.85ul / cm )

Green, 1/16x 0.75 mm ( 4.42ul / cm )

Orange, 1/16x 0.5 mm ( 1.96ul / cm )

Blue, 1/16x 0.25 mm ( 0.49ul / cm )

Teflon / Tefzel Tubing

3/16 OD

18-1112-47

1/8 OD

18-1121-16

1/16 OD

18-1119-74

Tubing Kit Changes - KTApurifier 10

GENERAL FLOW (at delivery)

G5
10

0u
l

Injection valve V1

G9
Outlet Valve V4 (7ul)

Accumulator
(40ul)

UV Flow cell

G8
G6
0.25 mm i.d
Blue PEEK
G5 400mm
G6 160mm
G7 80mm
G8 130mm
G9 500mm

G7
Conductivity
Flow cell (14ul)
FR-904
(0.2Mpa, 5ul)

NB: Never use

the FR-904 with
the pH electrode
pH Flow cell (100ul)

Tubing Kit
AKTApurifier 10 , 0.50 tubing kit
AKTApurifier 10 , 0.50 tubing kit + pH electrode
AKTApurifier 10 , 0.25 tubing kit

No Frac

Frac 901
232
354
88

Frac 950
382
504
238

Tubing Kit Changes - KTApurifier 10

HIGH FLOW

G5
10

0u
l

Injection valve V1

H9
Outlet Valve V4 (7ul)

Accumulator
(40ul)

UV Flow cell

H8
H6

H7
Conductivity
Flow cell (14ul)

0.50 mm i.d
Orange PEEK
H6 160mm
H7 80mm
H8 130mm
H9 500mm

FR-904
(0.2Mpa, 5ul)

NB: Never use

the FR-904 with
the pH electrode

pH Flow cell (100ul)

Tubing Kit
AKTApurifier 10 , 0.50 tubing kit
AKTApurifier 10 , 0.50 tubing kit + pH electrode
AKTApurifier 10 , 0.25 tubing kit

No Frac

Frac 901
232
354
88

Frac 950
382
504
238

Tubing Kit Changes - KTApurifier 100

HIGH FLOW (at delivery)

G5
10

0u
l

Injection valve V1

G9
Outlet Valve V4 (7ul)

Accumulator
(40ul)

UV Flow cell

G8
G6
0.75 mm i.d
Green PEEK
G6
G7
G8
G9

G7
Conductivity
Flow cell (14ul)
FR-904
(0.2Mpa, 5ul)

160mm
80mm
130mm
500mm

NB: Never use

the FR-904 with
the pH electrode
pH Flow cell (100ul)

Tubing Kit
AKTApurifier 100 , 0.50 tubing kit
AKTApurifier 100 , 0.75 tubing kit
AKTApurifier 100 , 1.00 tubing kit
AKTApurifier 100 , 0.50 tubing kit + pH electrode
AKTApurifier 100 , 0.75 tubing kit + pH electrode
AKTApurifier 100 , 1.00 tubing kit + pH electrode
AKTAbasic 100 , 0.75 tubing

No Frac

290

Frac 901
232
470
769
354
672
955

Frac 950
382
620
809
504
862
995

Tubing Kit Changes - KTApurifier 100

LOW FLOW

L5
10

0u
l

Injection valve V1

L9
Outlet Valve V4 (7ul)

Accumulator
(40ul)

UV Flow cell

L8
L6

0.50 mm i.d
Orange PEEK
L6
L7
L8
L9

L7
Conductivity
Flow cell (14ul)

160mm
80mm
130mm
500mm

FR-904
(0.2Mpa, 5ul)

NB: Never use

the FR-904 with
the pH electrode

pH Flow cell (100ul)

Tubing Kit
AKTApurifier
AKTApurifier
AKTApurifier
AKTApurifier
AKTApurifier
AKTApurifier

No Frac
100
100
100
100
100
100

,
,
,
,
,
,

0.50
0.75
1.00
0.50
0.75
1.00

tubing
tubing
tubing
tubing
tubing
tubing

kit
kit
kit
kit + pH electrode
kit + pH electrode
kit + pH electrode

Frac 901
232
470
769
354
672
955

Frac 950
382
620
809
504
862
995

A-900 Autosampler
INV-907
1

A-900
2

3
4

3
MANUAL
SAMPLE
LOAD

LOAD

6
5

Waste
COLUMN

Sample

PUMP

Waste
LOAD

Syringe

INV-907
1

A-900
2

1
6

3
4

3
MANUAL
SAMPLE
LOAD

INJECT
Syringe

Sample

2
4

6
5

Waste

Waste
COLUMN

PUMP

LOAD

Needle Height
Standard Vial
Standard Vial
+
Needle Height Insert
volume left

2mm
450ul

1mm
350ul

0mm
200ul

3mm
12ul

A-900

Sample Injection ( Flushed Loop Injection )

INJECT
INJECT

INJECT
INJECT

LOAD
LOAD
pump

waste

INJECT
INJECT

pump
waste

waste

waste
column

column

pump

pump

column

column

Sample needle
sample

INJECT,sample
sample
InInINJECT,
flushesthe
thevalve
valve
flushes
channel.
channel.
Min.( (30ul
30ul) )with
with
Min.
AirSegment
Segment
Air
Min.( (35ul
35ul) )without
without
Min.
Airsegment
segment
Air

sample

LOADThe
Thesample
sample
InInLOAD
drawnup
upto
to
isisdrawn
completely
completely
fillthe
theloop.
loop.
fill
noless
lessthan
thantwice
twice
( (no
loopvolume
volume
loop
recommended) )
recommended

Wash
Reservoir

sample

INJECTsample
sample
InInINJECT
washedonto
onto
isiswashed
thecolumn.
column.
the

sample

Finalinternal
internal
Final
needlewash.
wash.
needle
pump
waste
column

INJECT
INJECT

OptionalExternal
External
Optional
needlewash.
wash.
needle

sample
transport
fluid

A-900

Sample Injection ( Partial Loop Injection )

INJECT
INJECT

INJECT
INJECT

LOAD
LOAD

INJECT
INJECT
waste

pump

pump
waste

waste

sample

INJECT,sample
sample
InInINJECT,
flushesthe
thevalve
valve
flushes
channel.
channel.
Min.( (30ul
30ul) )with
with
Min.
AirSegment
Segment
Air
Min.( (35ul
35ul) )without
without
Min.
Airsegment
segment
Air

waste

waste

column

Sample needle

pump

pump

column

Sample needle

sample

Thesample
sampleisisdrawn
drawn
The
upto
topartially
partiallyfill
fillthe
the
up
loopwith
with
loop
programmedvolume.
volume.
programmed
nomore
morethan
thanhalf
half
( (no
loopvolume
volume
loop
recommended) )
recommended

column

column

Sample needle

sample

injectsample
sample
InIninject
washedonto
onto
isiswashed
thecolumn.
column.
the

Wash
Reservoir

sample

Finalinternal
internal
Final
needlewash.
wash.
needle
pump
waste
column

INJECT
INJECT

OptionalExternal
External
Optional
needlewash.
wash.
needle

sample
transport
fluid

A-900

Sample Injection ( ul Pick-up Injection )

INJECT
INJECT

LOAD
LOAD
pump

pump
waste

transport
fluid

sample

sample

Thesample
sampleisis
The
drawnup
upfrom
from
drawn
thesample
samplevial
vial
the

INJECT
INJECT

column

Sample needle

Sample needle
sample

sample

Thesample
sampleisis
The
washedinto
into
washed
theloop
loopwith
with
the
transportfluid
fluid
transport

pump

injectsample
sample
InIninject
washedonto
ontothe
theco
column
isiswashed
lumn
pump
waste

waste

Finalinternal
internal
Final
needlewash.
wash.
needle
Wash
Reservoir

pump

column
Sample needle

inject,valve
valve
InIninject,
flusheswith
with
flushes
transportfluid
fluid
transport

pump

column

column

INJECT
INJECT

LOAD
LOAD

column

sample

column

INJECT
INJECT

Optionalneedle
needle
Optional
wash.
wash.

sample
transport
fluid

System Compensation Volume

Equilibration Phase
A
B

COLUMN TOP
0%

A
B

COLUMN TOP
20%
Effect

8 mls

Command

20%B

0%B

8mls

Gradient Delay Volume

Gradient Phase

A
B

COLUMN TOP
0%

A
B

COLUMN TOP
100%
Effect

16 mls

100%B

Gradient Delay

Command 100%

Column Pressure ( compensation ) Flow Restrictor

Column Pressures

Column Hardware Pressure Limits

Column Shatters DANGEROUS

Column Matrix Pressure Limits

Matrix Collapses EXPENSIVE

0.3Reading
Reading
0.3

Atmos
Atmos
Press
Press

0.0MPa
MPa
0.0
Pre-column pressure
= 0 MPa

Pump
Pump
0.3Mpa
Mpa
0.3

0.3 MPa
GELPress
PressDrop
Drop
GEL

0.3MPa
MPa
0.3

(0.3- -0.0)
0.0)
(0.3

0.0 MPa

0.0MPa
MPa
0.0

0.5Reading
Reading
0.5
Pre-column pressure
= 0 MPa

Gel Top Pressure

Pump
Pump
0.3Mpa
Mpa
0.3

0.3 MPa

ColumnPress
PressDrop
Drop
Column

GELPress
PressDrop
Drop
GEL

Colu
mnPress
PressDrop
Drop
Colu
mn

(0.3++0.0)
0.0)
(0.3

(0.3- -0.2)
0.2)
(0.3

(0.5++0.2)
0.2)
(0.5

0.3MPa
MPa
0.3

Gel Bottom Pressure

0.1MPa
MPa
0.1

0.2 MPa

0.2MPa
MPa
0.2
FlowRestrictor
Restrictor
Flow

0.5MPa
MPa
0.5