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Coba e 411
Coba e 411
Compendium of
Background Information
COBI-CD
cobas e 411
Revision history
Intended use This CD is provided as an information source for background information regarding
the cobas e 411 analyzer The information on this CD is available in PDF-format and
requires Adobe Acrobat Reader to be installed.
Trademarks COBAS, COBAS C, COBAS E, ELECSYS, and LIFE NEEDS ANSWERS are
trademarks of Roche.
All other trademarks are the propery of their respective owners.
Instrument approvals The cobas e 411 analyzer meets the requirements stated in Directive 98/79/EC of the
European Parliament and the Council of the European Union (EU) on in vitro
diagnostic medical devices. Furthermore, the cobas e 411 analyzer is manufactured
and tested according to International Standard IEC 61010-1, 2nd edition, “Safety
requirements for electrical equipment for measurement, control and laboratory use,
Part 1: General requirements”. This International Standard is equivalent to the
national standards Underwriters Laboratories (UL) 61010-1 2nd edition for the
USA, and CAN/CSA C22.2 No. 61010-1:2004 for Canada. Compliance is
demonstrated by the following marks:
C US Issued by Underwriters Laboratories, Inc. (UL) for Canada and the USA.
®
Notice to the purchaser The purchase of this product allows the purchaser to use it solely for detection by ECL
Technology for human in vitro diagnostic uses. No general patent or other license
of any kind other than this specific right of use from purchase is granted hereby.
This product may not be used by purchaser to conduct life science research and/or
Roche Diagnostics
Contact addresses
Manufacturer
Hitachi High-Technologies Corporation
24-14. Nishi-shimbashi. 1-chome. Minato-ku
Tokyo. 105-8717 JAPAN
Authorized Representative
Roche Diagnostics GmbH
Sandhofer Strasse 116
D-68305 Mannheim
Germany
Conventions used
Visual cues are used to help you quickly locate and interpret information in this
manual. This section explains formatting conventions used in this manual.
Caution
Warning
Laser Radiation
Biohazard
Abbreviation Definition
A
ANSI American National Standards Institute
C
CBT Computer Based Training
CCITT Comité consultatif international téléphonique et télégraphique
(Consultative Committee on International Telegraph and
Telephone)
CE Conformité Européenne’
CLAS 2 Clinical Laboratory Automation System 2
CLIA Clinical Laboratory Improvement Amendments
COBI-CD Compendium of Background Information
CSA Canadian Standards Association
Abbreviation Definition
D
dBA decibel weighted against the A-frequency response curve. This curve
approximates the audible range of the human ear.
DIL diluent
E
EC European Community
ECL electrochemiluminescence
EMC electromagnetic compatibility
EN european standard
F
FIFO first in first out
H
HCFA Health Care Financing Administration
I
IEC International Electrical Commission
IS internal standard (ISE module)
IVD in vitro diagnostic directive
K
KVA kilovolt-Ampere. Unit for expressing rating of AC electrical
machinery.
L
LDL lower detection limit see analytical sensitivity
LIS laboratory information system
LLD liquid level detection
M
MSDS material safety data sheet
N
NCCLS National Committee for Clinical Laboratory Standards
P
PC/CC ProCell/CleanCell
Q
QC quality control
R
REF reference solution for ISE module
S
SD standard deviation
S/R sample/reagent
SVGA Super Video Graphics Adapter
T
TPA tripropylamine
U
UL Underwriters Laboratories Inc.
Abbreviation Definition
V
VDE Verband Deutscher Elektrotechniker (association of
German electrical engineers)
Table of contents
Revision history 2
Contact addresses 3
Quality control Part D
Conventions used 4
Table of contents 7 5 Quality control concept
Control target value (first) assignment D-5
Mechanical theory Part A
Index Part E
1 Mechanical theory
Introduction A-5 Index E-3
Preparative operations A-6
Test protocols A-7
Assay sequence A-8
Workflow and throughput A-11
Operation flow in analysis A-13
Detailed assay sequence A-14
Dilution steps A-21
Pretreatment steps A-22
Analyzer status conditions A-23
2 ECL technology
ECL measuring principles B-5
Advantages of ECL technology B-10
3 Test principles
Test principles C-5
4 Reagent concept
Introduction C-15
Data transfer media C-15
Data transfer rules C-16
Reagents for cobas e 411 analyzer tests C-16
Product labeling C-18
Data links C-19
Calibration C-21
Master calibration C-22
Lot calibration C-23
Reagent pack calibration C-23
Difference between lot and reagent calibration C-24
Calibration procedures C-25
Calibration stability C-26
Calibration validation C-26
Calibration assessment C-27
Calibration of quantitative assays C-30
Calibration of qualitative assays C-33
Result calculation for qualitative assays C-33
Mechanical theory A
1 Mechanical theory........................................................................A-3
cobas e 411 1 Mechanical theory
Table of contents
Mechanical theory
This chap
provides
overview
mechanic
theory o
cobas e 4
analyzer.
assay seq
and oper
flow are
described
well as d
steps.
1 In this
chapte
Chapt
Introduc
..............
31
Preparati
operatio
..............
32
Test pro
..............
33
Assay
sequenc
..............
34
Run ope
..............
34
First incu
°C
..............
34
Pipetting
reagent
Roche Diagnostics
COBI-CD · Version 1.0 A-3
........................................................................................................................ put
Seco ..............
nd 37
incu 9-minute
batio ..............
n at 37
37 °C 18-minu
........................................................................................................................ ..............
Pipe 37
tting Combina
of and 18-m
addi ..............
tiona 37
l 27-minu
reag ..............
ent 38
(pret Combina
reat and 27-m
ment ..............
assay 38
s).....................................................................................................................
Typical t
Third duration
incu ..............
batio 38
n at
Operatio
37 °C
in analys
(pret
..............
reat
39
ment
Detailed
assay
sequenc
s).....................................................................................................................
..............
Aspir
40
ating
Preopera
the
steps
reac
..............
tion
40
mixt
ure Dispensa
........................................................................................................................ reagent
2, and sa
Clea
(disk sys
ning
..............
the
41
meas
uring Dispensa
cell reagent
........................................................................................................................ 2, and sa
(rack sys
Finali
..............
zatio
43
n......................................................................................................................
First incu
Workflow
..............
and
44
throughput
.................................................................................................................................... Microbe
preparati
Effects
..............
of test
45
combina
tions on
through
cobas e 411 1 Mechanical theory
Table of contents
Roche Diagnostics
COBI-CD · Version 1.0 A-4
cobas e 411 1 Mechanical theory
Introduction
Roche Diagnostics
COBI-CD · Version 1.0 A-5
Preparative operations
Preparative operations
Once the analyzer is switched on, the initialization process starts. During
initialization, the mechanisms are reset to their home positions.
e Figure A-1 shows the run preparation process for the cobas e 411 hardware.
Start
First order?
Were reagents
exchanged? No
Yes Counting
AssayTips and AssayCups
Scan of the reagent barcode
Volume check for ProCell and
CleanCell
Have 90 or Clearing
more the incubator and the AssayTip and AssayCup
No
minutes passed since the last mixing? trays
Yes
Microbeads mixing
Enough
Scheduling Preparation cycle
Resume cycle
Pipetting continues
Sipping continues
Test
protocols Test protocols
There are 28 test protocols that can be used on the analyzer. These protocols are
predefined by Roche Diagnostics for each test and cannot be changed by the operator.
Assay sequence
Run operation
After the appropriate test selections for patient samples are made in the software,
operation is started according to the predetermined test protocol for each assay
selected. Initially, at least one reagent (R1 or R2) and the sample or microbeads
(M) are aspirated one after another by the S/R probe. After each aspiration, the
outside of the S/R probe AssayTip is cleaned at the rinse station. The sample and
reagents are dispensed into a new AssayCup and the AssayTip is ejected into the solid
waste tray.
For some tests that require sample dilution or pretreatment, diluent or pretreatment
reagent is pipetted together with sample into an AssayCup. An aliquot of the diluted/
pretreated sample is then dispensed with reagent into a second AssayCup. Therefore,
certain tests with predilution/pretreatment may require two or more AssayCups.
e For more information on dilution, see Dilution steps on page A-21.
First incubation at 37
°C
The incubation times are 4.5 or 9 minutes long, depending on the test. Some tests
require only two incubation periods, whereas tests that include pretreatment can
require three incubation periods. During the incubation step(s) the immune
complex products are formed.
Pipetting of additional
reagent
Some assays (usually those with more than one incubation step) require additional
reagent pipetting. As in the initial reagent pipetting step, a new AssayTip is picked up
before reagent aspiration. The S/R probe AssayTip is washed at the rinse station after
each liquid aspiration. The liquid is then dispensed into the corresponding AssayCup
where the sample and other liquids were dispensed in the first pipetting step. The
probe rises while dispensing the reaction mixture back into the AssayCup, thereby
mixing the solution and accelerating the reaction in the AssayCup. The AssayTip is
discarded into the solid waste tray when pipetting is complete.
Roche Diagnostics
Second incubation at 37 °C
Finalization
Thirty minutes after documentation of the last result, the sipper pipetter flushes
system water through the sipper probe, and then fills the measuring cell with
ProCell before the analyzer returns to Standby mode.
After this procedure, every 30 minutes the waste pump of the S/R rinse station runs
for 2 seconds (waste consumption approximately 12 mL). This procedure stops if
the operation switch is switched off.
e Figure A-2 shows the finalization process for the cobas e 411 hardware.
Last sipping
Finalization
Sipper prime
Standby
The various available tests have different durations. The throughput of the cobas e
411 analyzer depends upon the way in which tests of a given duration are
combined, as explained for each of the following combinations. There may be short
periods of throughput slow-down on the disk system due to the loading of a new
sample disk. Such gaps do not occur when the rack system is used, because the
Roche Diagnostics/ Hitachi 5-position racks load continuously.
18-minute tests have two incubation periods, each of 9 minutes duration. If only
18- minute tests are performed, the optimal throughput will always be reached
regardless of the test mixture.
All 18-minute tests follow the same time protocol. Therefore, there will be no
timing conflicts. In one 42-second cycle, the cobas e 411 will simultaneously perform
S1 (first reagent pipetting), S2 (second reagent pipetting), and D (detection).
27-minute tests have three incubation periods, each of 9 minutes duration. If only 27-
minute assays are performed, the throughput of the cobas e 411 is reduced to 44
results per hour. Every 13 cycles, the cobas e 411 comes into a timing problem. It is
not possible to perform a S0 (pretreatment pipetting) together with a S1 (first reagent
pipetting) within one 42 second cycle. When this happens, the instrument will
stand for 13 cycles (9 minutes) until it can pipette again without conflict.
e Table A-2 contains details of the duration of some typical tests. This is not a complete list
of tests, but is provided as an example.
Roche Diagnostics
Pre-start inspection
Routine operation
Calibration and control
Rerun Routine or STAT(a) sampling
Pre-routine operation
Assigned
Results
Sampling Stop
(Finalization, Stop, and Standby)
Maintenance
Switch off
(a)
Short Turn Around Time
Figure A-3 Operational process
1 Mechanical theory cobas e 411
Detailed assay sequence
The mechanical process of the instrument is described below with a sandwich test,
TSH (thyroid-stimulating hormone), used as an example. This example assumes
that the reagent pack was already registered by the analyzer and does not need
calibration. All results are calculated on the basis of an existing lot calibration.
Preoperational steps
When Start is pressed from Standby mode, the following preoperational steps occur:
1. The analyzer resets all mechanisms to their respective home positions and accesses
the data disk. Next, the S/R pipetter primes the S/R probe.
2. The gripper checks for an AssayTip in position number 1 of the AssayTip trays. If
this position is empty, the gripper remembers where it last left off and checks that
position. If this position is empty, the gripper considers the whole tray empty and
the Inventory screen is updated accordingly.
If the analyzer is in S. Stop, the gripper remembers where it last left off and checks for an
AssayTip in that position.
3. During the AssayTip check, the S/R probe is checked for the presence of
an AssayTip. The probe moves to the AssayTip eject station and performs
the movements to eject an AssayTip. If an AssayTip is present, it is
ejected.
4. After the AssayTip check is complete, the AssayCups are checked in the same
manner. During the cup check, the analyzer finishes priming the probes.
5. Next, the gripper checks the last three of the five positions on the pipetting
station. If an AssayCup is present, the analyzer goes through the following
cup disposal sequence:
a) The gripper places an AssayTip in position 1 of the pipetting station.
b) The S/R probe picks up the AssayTip, descends into the AssayCup,
and aspirates any liquid.
c) The AssayCup is discarded, while the S/R probe moves to the rinse station and
dispenses any aspirated liquid.
d) The AssayTip is washed and then discarded.
6. The gripper moves to the incubator, where it checks all 32 incubator positions.
If an AssayCup is present, the gripper moves the AssayCup to position 5 on the
pipetting station and uses the same procedure listed in step 5 to discard the
AssayCup.
7. The S/R probe AssayTip is discarded after all the incubator positions are checked.
Roche Diagnostics
5. The S/R probe moves from its Standby position to the R1 aspiration position.
While activating liquid level detection, the probe descends until it is 2 mm
below the reagent surface and aspirates 50 µL of R1.
While the probe is aspirating R1, the gripper puts another AssayTip in position 1
of the pipetting station.
6. If the S/R probe does not detect liquid as it descends, no reagent aspiration can
occur, and an alarm is generated.
7. After R1 aspiration, the S/R probe rises and moves to the rinse station. To prevent
the aspirated R1 from coming into contact with the water in the rinse station,
the probe aspirates 10 µL of air. The rinse station externally washes the
AssayTip.
8. During step 7, the reagent rotor rotates until the TSH reagent pack is in the
R2 position.
9. The S/R probe moves from the rinse station to the R2 aspiration position while
aspirating another 10 µL of air. This air layer prevents R1 from mixing with R2.
While activating liquid level detection, the probe descends until it is 2 mm
below the reagent surface and aspirates 50 µL of R2. While the probe is aspirating
R2, the gripper moves an AssayCup to position 5 of the pipetting station.
e See Figure A-4 for the location of the R2 aspiration position.
10. Upon completion of R2 aspiration, the S/R probe rises and moves to the rinse
station. To prevent the aspirated R2 from coming into contact with the water in
the rinse station, the probe aspirates another 10 µL of air. The rinse station
externally washes the AssayTip.
11. After R2 aspiration, the reagent rotor rotates until the TSH reagent pack is at
the cap open/close mechanism. The mechanism moves out and closes the
caps.
12. The S/R probe moves from the rinse station to the sampling position while
aspirating another 10 µL of air. While activating liquid level detection, the
probe descends until it is 2 mm below the sample surface and aspirates 50 µL of
sample. During sample aspiration, clot detection is activated.
A
A Sampling position
Figure A-5 Sampling position (disk system)
13. The S/R probe moves from the sampling position to position 5 of the pipetting
station. The probe descends until the tip reaches 2 mm below the calculated level
of the reaction mixture surface and dispenses the sample, R2, and R1. The probe's
downward displacement is determined by calculating the volume of the reaction
mixture for the sample and using downward-displacement tables in the software.
The probe does not rise during dispensation.
e See Figure A-5 for the location of the sampling position for disk systems.
14. After dispensation, the S/R probe moves to the tip eject position and ejects the
AssayTip.
Detailed assay sequence
2. The pusher arm pushes the racks in the A-Line forward to the B-Line. The arm
returns to its home position. The first rack loads on the B-Line.
e For additional information on the A-Line and B-Line, see the Sample/reagent area
components section in the Analyzer components chapter of the cobas e 411 analyzer
Operator’s Manual.
3. As the rack incrementally moves on the B-Line, the rack barcode reader scans
all five rack positions and rack ID. When scanning is complete, position 1 of the
rack is in the sampling position.
4. The S/R probe moves to position 1 of the pipetting station, descends to obtain the
AssayTip, rises, and returns to its Standby position.
5. During this time, the reagent rotor rotates until the TSH reagent pack is at the
cap-open/close mechanism. The mechanism moves forward and opens the caps
on the reagent pack. The disk rotates again to move the TSH reagent to the
R1 position.
e See Figure A-4 on page A-15 for details of the R1 and R2 aspiration positions.
6. The S/R probe moves from its Standby position to the R1 aspiration position.
While activating liquid level detection, the probe descends until it is 2 mm
below the surface of the reagent and aspirates 50 µL of R1.
e See Figure A-4 for the location of the R1 aspiration position.
While aspirating R1, the gripper puts another AssayTip in position 1 of the
pipetting station.
7. If the S/R probe does not detect liquid during descent, no reagent aspiration can
occur, an alarm is generated.
8. After R1 aspiration, the S/R probe rises and moves to the rinse station. To prevent
the aspirated R1 from contacting the water in the rinse station, the probe aspirates
10 µL of air. The rinse station externally washes the AssayTip.
9. During step 8, the reagent rotor rotates until the TSH reagent pack is in the
R2 position.
10. The S/R probe moves from the rinse station to the R2 position while aspirating
another 10 µL of air. This air layer prevents R1 from mixing with R2. While
activating liquid level detection, the probe descends until it is 2 mm below the
surface of the reagent and aspirates 50 µL of R2. While aspirating R2 the
gripper moves an AssayCup to position 5 of the pipetting station.
e See Figure A-4 for the location of the R2 aspiration position.
11. Upon completion of R2 aspiration, the S/R probe rises and moves to the rinse
station. To prevent the aspirated R2 from coming into contact with the water in
the rinse station, the probe aspirates another 10 µL of air. The rinse station
externally washes the AssayTip.
12. After R2 aspiration, the reagent rotor rotates until the TSH reagent pack is at
the cap-open/close mechanism. The mechanism moves out and closes the
caps.
13. The S/R probe moves from the rinse station to the sampling position while
aspirating another 10 µL of air. While activating liquid level detection, the
probe
1 Mechanical theory cobas e 411
Detailed assay sequence
A Sampling position
Figure A-6 Sampling position (rack system)
14. The S/R probe moves from the sampling position to position 5 of the pipetting
station. The probe descends until the tip reaches 2 mm below where the calculated
level of the reaction mixture surface should be and dispenses the sample, R2, and
R1. The probe's downward displacement is determined by calculating the volume
of the reaction mixture for the sample and using downward-displacement tables
in the software. The probe does not rise during dispensation.
e See Figure A-6 for the location of the sampling position for rack systems.
15. After dispensation, the S/R probe moves to the tip eject position and ejects the
AssayTip.
First incubation
1. The gripper picks and transports the cup containing the reaction mixture
from the pipetting station to the incubator.
2. The cup is incubated at 37 °C for 9 minutes.
3. During incubation, the analyzer continues to perform operations for other
test(s) or sample(s), if necessary.
Roche Diagnostics
Microbead preparation
Before the first incubation is completed, the TSH microbeads are mixed to facilitate
their aspiration and dispensation.
1. The reagent rotor rotates until the TSH reagent pack is at the reagent cap-
open/ close mechanism. The mechanism moves out and opens the cap. The
disk moves the reagent pack to the mixing position.
2. The mixer moves over the reagent rotor and descends into the microbeads to 1.4
mm above the bottom of the bottle.
3. The mixer stirs the microbeads to obtain a homogeneous suspension.
4. During the mixing, the gripper obtains a fresh AssayTip and transports it
to position 2 of the pipetting station.
5. When mixing is complete, the mixer rises and returns to the rinse station where it
descends and rotates in the rinse station for washing.
6. At the same time, the reagent rotor rotates the TSH reagent pack to the microbead
pipetting position.
Second incubation
1. The gripper picks the cup containing the mixed reaction mixture and returns it
to the incubator.
2. The cup is incubated at 37 °C for 9 minutes.
3. During incubation, the analyzer continues to perform operations for other
test(s) or sample(s), if necessary.
Preparations for the measurement process
Before the second incubation is completed, the sipper probe aspirates ProCell into the
measuring cell to facilitate measurement.
1. The sipper probe moves from its home position to a ProCell bottle and
descends to 2 mm below the solution level and aspirates ProCell into the
measuring cell. As the probe descends, liquid level detection is activated. The
sipper probe can descend as low as 1.3 mm above the bottom of the ProCell
bottle.
2. The sipper probe rises.
Measurement process
1. The gripper picks and transports the cup that has completed its second incubation
from the incubator to the aspiration station.
2. The sipper probe moves to the aspiration station and descends into the cup.
3. When the sipper probe detects the reaction mixture in the cup, it aspirates 150 µL.
4. After aspiration, the sipper probe rises, aspirates 10 µL of air, and moves to the
sipper rinse station to descend for rinsing.
5. The gripper grasps the cup from the aspiration station, transports it to the
cup disposal opening, and discards the cup.
6. The sipper probe is rinsed.
7. The sipper probe rises and moves to the ProCell position, descends into the bottle
and aspirates ProCell in a set aspiration/dispensation sequence. The immune
complexes are magnetically captured on the working electrode, but unbound
reagent and sample are washed away by ProCell.
e For additional information on the measuring cell, see Chapter 2 ECL technology.
8. After the bound-free separation, a voltage is applied between the working
electrode and the counter electrode. The ECL reaction is initiated and measured
by the photomultiplier.
e For additional information on binding and bound-free separation, see Chapter 3 Test
principles.
9. The sipper probe rises and moves to the CleanCell position and aspirates 20 µL
of air. The probe then descends into the CleanCell bottle and aspirates reagent.
This procedure is repeated eight times. The alternatating flow of air and
cleaning solution washes the measuring cell. During this washing process, a
voltage is applied between the electrodes, which aids in the cleaning process.
10. The sipper probe moves to the sipper rinse station, aspirates 20 µL of air, and
descends into the rinse station for washing.
11. Finally, the sipper probe rises and moves to the ProCell bottle. The probe
descends into the bottle and aspirates 500 µL of ProCell. Next, the probe aspirates
90 µL of ProCell and moves to the rinse station. At the rinse station, the probe
dispenses 35 µL to flush the probe and prepare it for the next sample. During
the aspirations of the ProCell, a sequence of voltages is applied three times to
prepare the electrodes for the next measurement.
One cycle of the measurement process consumes approximately 2 mL each of ProCell
and CleanCell.
Dilution steps
There are certain analyzer functions that occur automatically while the analyzer is
switched on.
o While the analyzer is in operation, the solid waste tray periodically shakes for
1.5 seconds.
o While the analyzer is in Standby, the reagent rotor turns 90° every 30 minutes.
o While the analyzer is in Standby, the rinse stations for the S/R probe and
sipper probe are switched on for 2 seconds every 30 minutes.
o Microbeads undergo a long mix when starting from Standby and then every
90 minutes until pipetting starts.
o Microbeads undergo a short mix and then a short mix every 60 minutes for
each reagent pack.
Dilution steps
(1:50, 1:100)
Pretreatment steps
Pretreatment assay
The cobas e 411 analyzer has a number of status conditions. The status conditions
usually seen during routine operation or maintenance procedures are listed below.
Several other status conditions, most them seen during various adjustment or
maintenance procedures performed by a Roche Diagnostics service representative, are
Roche Diagnostics not included below.
A-22 COBI-CD · Version 1.0
cobas e 411 1 Mechanical theory
e Refer to the Alarm screen for further information about instrument alarms.
The analyzer is no longer able to continue operation. An alarm was issued. Take the
appropriate measures to resolve the problem.
The analyzer is already in A. Stop status when the lines stop operation.
The analyzer is already in A. Stop status when the A-Line stops supplying racks to
the B-Line.
BC card scan
This status is seen when a barcode card scan is initiated from the Control Definition
or Calibration Data screens.
An emergency stop condition exists. An alarm was issued. Take the appropriate
measures to resolve the problem.
Finalization
This is the status of the analyzer when it is between the status conditions S. Stop and
Standby.
Roche Diagnostics
COBI-CD · Version 1.0 A-23
Analyzer status conditions
Finalization maint.
Initialization
This status is seen when the cobas e 411 analyzer is switched on, or when Start is
pressed from Standby.
L. and A. reset all status occurs when the corresponding function is initiated from the
Maintenance screen. This function resets the analyzer and the lines.
All lines stop operation. An alarm was issued. Take the appropriate measures to
resolve the problem.
Liquid flow cleaning occurs when this function is initiated from the Maintenance
screen.
M. Cell preparation
Measuring cell (M. Cell) preparation occurs when this function is initiated from the
Maintenance screen.
Operation
This is the status during which the cobas e 411 analyzer performs its routine
operations.
A partial stop condition exists. An alarm was issued. Take the appropriate measures
to resolve the problem.
Analyzer status conditions
This status occurs when there are no more racks to process on the A-Line or B-Line.
Rack clear
Rack clear status occurs when the corresponding function is initiated from the
Maintenance screen. This function clears any remaining racks on the A-, B- or C-
Lines.
Reagent scan
This status is seen when a reagent scan is initiated from the Inventory screen.
This status occurs when the S/R (sample/reagent) pipetter prime is initiated from the
Maintenance screen.
This status is seen when the analyzer is monitoring the liquid level detection voltage
of the S/R (sample/reagent) probe. The check is initiated from the Voltage Monitor
screen (Utility) folder.
S. Stop-S. Scan
The analyzer is in S. Stop and a sample scan is requested from the Status screen, or S is
pressed while the analyzer is in S. Stop.
Sample scan
This status occurs when a sample scan is initiated from the Status screen.
Analyzer status conditions
The analyzer is monitoring the liquid level detection voltage of the sipper probe.
The check is initiated from the Voltage Monitor screen (Utility) folder.
This status occurs when the sipper pipetter prime is initiated from the Maintenance
screen.
Standby
Stop
This status occurs when Stop is pressed or when a Stop alarm condition exists. If
an alarm exists, take the appropriate measures to resolve the problem.
System reset
2 ECL technology....................................................................B-3
cobas e 411 2 ECL technology
Table of contents
ECL technology
This chap
provides
overview
the
electroch
minescen
technolo
the coba
analyzer
system. T
use of th
rutheniu
complex
the meas
cell in wh
the reac
occurs a
describe
2 In this
chapte
Chapt
ECL
measuri
principle
.............
5
Use of th
rutheniu
complex
..............
5
The ECL
at the el
surface
..............
6
ECL sign
generati
.............
Roche Diagnostics 8
COBI-CD · Version 1.0 B-3
E
C
L
m
e
a
s
u
r
i
n
g
c
e
l
l
........................................................................................................................
9
Adv
anta
ges
of
ECL
tech
nolo
gy
....................................................................................................................................
cobas e 411 2 ECL technology
Table of contents
Roche Diagnostics
COBI-CD · Version 1.0 B-4
cobas e 411 2 ECL technology
ECL measuring principles
The chemiluminescent reactions that lead to the emission of light from the
ruthenium complex are triggered electrically, rather than chemically. This is achieved
by applying a voltage to the immunological complexes (including the ruthenium
complex) that are attached to streptavidin-coated microbeads. The advantage of
electrically initiating the chemiluminescent reaction is that the entire reaction can be
precisely controlled.
ECL technology uses a ruthenium chelate as the complex for the development of
light. Salts of ruthenium-tris(bipyridyl) are stable, water-soluble compounds. The
bipyridyl ligands can be readily modified with reactive groups to form activated
chemiluminescent compounds.
For the development of ECL immunoassays, a N-hydroxysuccinimide (NHS) ester of
a modified Ru(bpy)3 complex is used because it can be easily coupled with amino
groups of proteins, haptens, and nucleic acids. This allows the detection technology
to be applied to a wide variety of analytes.
2+
O
O
N N
N N O
O
Ru
N N
Roche Diagnostics
COBI-CD · Version 1.0 B-5
ECL measuring principles
Photon
Diffusion
Magnetic
TPA• microbead
-H+
TPA TPA•+
e-
e-Electrode
In turn, the ruthenium complex also releases an electron at the surface of the
electrode thus oxidizing to form the Ru(bpy)33+ cation. This ruthenium cation is the
second reaction component for the following chemiluminescent reaction with the
TPA radical.
e For further information on the ECL reaction at the electrode surface, refer to Figure B-3.
Electrode surface
TPA•+
e- -H+
TPA
- TPA•
Ru(bpy) 3+e
3
e-
2+
Ru(bpy)3
Ru(bpy) 2+ excited state
groun 3
d
state Photon
(620 nm)
TPAo and Ru(bpy)33+ react with one another, whereby Ru(bpy)33+ is reduced to
Ru(bpy)32+ and at the same time forms an excited state through energy transfer.
This
excited state is unstable and decays, with emission of a photon at 620 nm to its
original state. The reaction cycle then starts again. The tripropylamine radical reduces
to by-products that do not affect the chemiluminescence process. TPA is used up and
therefore must be present in excess. The reaction is controlled by diffusion of the TPA
and the amount of ruthenium complex present. As TPA in the electrical field is
depleted, the signal strength (light) is slowly reduced once the maximum is
reached.
Although TPA is depleted during measurement, the ruthenium ground-state
complex is continually regenerated. This means that the ruthenium complex can
perform many light-generating cycles during the measurement process. This has an
inherent amplification effect that contributes to the sensitivity of the technology.
Many photons can be created from one antigen-antibody complex.
ECL measuring principles
The following figure illustrates a typical ECL signal generation. Viewed from an
electrical perspective, the reaction can be explained as follows: When a voltage is
applied to the electrode of the measuring cell, a brief peak of light emission occurs,
which can be detected as the resulting ECL signal. A defined area under the curve is
measured around the intensity maximum.
350,000
300,000 1200
250,000
200,000 900
150,000
100,000
50,000 600
0
0.00
300
The dotted line indicates the voltage at the electrode used to generate the ECL signal.
The solid line is the actual light output measured by the photomultiplier detector.
ECL measuring cell
The core of the detection unit is the ECL measuring cell, which is designed as a flow-
through cell. The following figure shows the main components of the measuring cell:
A B C D E
H
I
J
O
N M L K
A Screw B Counter electrode C Optical window
D Distance washer E Top cell F Cell gap
G Gasket H O-ring I Diaphram
J Reference electrode K Outlet fitting L Working electrode
M Movable magnet N Inlet fitting O Cell body
The temperature is maintained at 28°C . Three operating steps are performed in the
measuring cell:
o Bound/free separation
Using a magnet, the streptavidin microbeads that are coated with antigen-
antibody complexes are uniformly deposited on the working electrode. A system
buffer (ProCell) is used to wash the particles on the working electrode and to flush
out the excess reagent and sample materials from the measuring cell.
o ECL reaction
To initiate the ECL reaction, the magnet is removed and a voltage is applied to the
electrode. The microbeads that are coated with antigen-antibody complexes are
deposited onto the electrode. The light emission is measured with a
Advantages of ECL technology
photomultiplier. The system then uses the corresponding signals for the
calculation of results.
o Release of microbeads and cell cleaning
Once the measurement is completed, the paramagnetic microbeads are washed
away from the electrode surface with a special cleaning solution (CleanCell).
The surface of the measuring cell is regenerated by varying the potential on the
electrode. The cell is then ready for another measurement.
B C
A D
F
G E
A Magnetic microbeads with bound antigen- B Photomultiplier
antibody complex
C Counter electrode D Unbound antibody (ruthenium-labeled)
E Flow channel F Magnet
G Working electrode
3 Test principles...............................................................................C-3
4 Reagent concept . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . C-13
cobas e 411 3 Test principles
Table of contents
Test principles
This chap
provides
overview
the
immuno
test prin
used by
cobas e 4
analyzer.
3 In this
chapte
Chapt
Test prin
..............
13
Competi
principle
..............
14
Sandwic
principle
..............
16
Bridging
..............
18
Roche Diagnostics
COBI-CD · Version 1.0 C-3
cobas e 411 3 Test principles
Table of contents
Roche Diagnostics
COBI-CD · Version 1.0 C-4
cobas e 411 3 Test principles
Test principles
Test principles
Streptavidin-biotin binding
Roche Diagnostics
COBI-CD · Version 1.0 C-5
Test principles
Competitive principle
FIRST REACTION
SECOND REACTION
LIGHT REACTION
Signal (light)
TPA ECL
Ruthenium-labeled antibody
Antigen
Biotinylated antigen
Streptavidin-coated microbead TPA Tripropylamine
Sandwich principle
FIRST REACTION
Serum constituents
SECOND REACTION
LIGHT REACTION
Signal (light)
TPA ECL
Bridging principle
The bridging principle is similar to the sandwich principle, except that the assay is
designed to detect antibodies (for example, IgG, IgM, and IgA), not antigens. This is
accomplished by including biotinylated and ruthenium-labeled antigens in the
reagents for which the targeted antibody has affinity.
e Refer to Figure C-4 on page C-11 for an illustration of the bridging principle.
o In the first step, serum antibodies bind with the biotinylated and
ruthenium- labeled antigens to form an immune complex.
o The immune complex then reacts with streptavidin-coated microbeads
through the action of the biotinylated antigen.
o After the second incubation, the reaction mixture containing the immune
complexes is transported into the measuring cell; the immune complexes are
magnetically entrapped on the working electrode, and the unbound reagent and
sample are washed away by ProCell.
o In the ECL reaction, the conjugate is a ruthenium based derivative and the
chemiluminescent reaction is electrically stimulated to produce light. The amount
of light produced is directly proportional to the amount of analyte in the
sample.
The concentration of the antibody is evaluated and calculated by means of a
calibration curve that was established using standards of known antibody
concentrations.
Roche Diagnostics
FIRST REACTION
Serum
constituents
SECOND REACTION
LIGHT REACTION
Signal (light)
TPA ECL
Concentration
Magnetic force and electrical potential
Biotinylated antigen
Ruthenium- labeled antigen
Reagent concept
This chap
provides
overview
types of
reagents
on the co
411 analy
system. I
describes
various r
containe
used, and
provides
overview
system-r
reagent
managem
explainin
processe
as how th
system
registers
reagents
how it
monitors
reagent
consump
4 In this
chapte
Chapt
Introduc
..............
15
Data tran
media
..............
15
Data tran
rules
Roche Diagnostics
COBI-CD · Version 1.0 C-13
.................................................................................................................................... stability
Reagent ..............
s for 26
cobas e Calibrati
411 validatio
analyze ..............
r tests 26
.................................................................................................................................... Calibrati
Dilu assessm
ents ..............
.............................................................................................................................. 27
Syst Missing
em ..............
reag 27
ents Monoton
.............................................................................................................................. curve
Cali (quantita
brat assays o
ors ..............
and 27
cont Slope (qu
rols assays o
.............................................................................................................................. ..............
Rea 27
gent Calibrati
pac (quantita
ks.......................................................................................................................... assays o
Product ..............
labeling 28
.................................................................................................................................... Minimum
Data ..............
links............................................................................................................................ 29
Calibra Minimum
tion............................................................................................................................. um signa
Master (qualitati
calibrati only)
on............................................................................................................................... ..............
Lot 29
calibrati Minimum
on............................................................................................................................... differenc
Reagen (quantita
t pack assays o
calibrati ..............
on............................................................................................................................... 29
Differen Minimum
ce acceptab
betwee differenc
n lot (qualitati
and only)
reagent ..............
calibrati 29
on...............................................................................................................................
Calibra
tion
proced
ures.............................................................................................................................
Calibra
tion
cobas e 411 4 Reagent concept
Table of contents
Deviation of duplication.......................................................................................29
System errors.......................................................................................................29
Cal.........................................................................................................................30
Target (quantitative assays only).........................................................................30
Cutoff (qualitative assays only)............................................................................30
Borderline (qualitative assays only).....................................................................30
Calibration of quantitative assays..............................................................................30
Rodbard function.................................................................................................31
Linear calibration function...................................................................................32
Linear reciprocal calibration function..................................................................32
Calibration of qualitative assays................................................................................33
Result calculation for qualitative assays....................................................................33
Roche Diagnostics
COBI-CD · Version 1.0 C-14
cobas e 411 4 Reagent concept
Introduction
Introduction Elecsys r
packs (co
packs) h
special 2
dimensio
barcode
allows fu
automati
registrati
managem
reagent
informati
advantag
barcode
manual e
addition
monitori
necessar
ready-to-
reagents
loaded in
the 18 po
the reage
Reagents
available
analysis a
barcodes
scanned.
The hand
calibrato
Roche D
controls
to that o
reagents
calibrato
supplied
use. Lyop
controls
calibrato
be prepa
transferr
the appr
containe
quantitati
assays, c
and cont
informati
stored on
barcode
qualitati
all inform
necessar
calibratio
encoded
barcode
Roche Diagnostics
COBI-CD · Version 1.0 C-15
4 Reagent concept cobas e 411
Data Barcode
calibrato
transfer controls
carry inf
media such as c
or contro
identifica
The level num
following lot ID.
data sources
Reagent
are available
barcodes
for cobas e
downloa
411 analyzer
encode a
applications: informati
1. Barcode including
s on applicati
reagent calibratio
packs validatio
(2D and expi
matrix e For furt
barcode) infor
o Reagent infor
pack enco
o Diluent the b
reagent label
pack Data
page
o Pretreatmen
t reagent
pack
o BlankCell
2. Barcode
s on
calibrato
r and
controls
vials (1D
barcode)
o Calibrator
primary vial
o Control
primary vial
3. cobas
Link and
system
database
(created
during
installati
on)
o Assay
o Calibrator
and barcode
card
o Control and
barcode
card
Roche Diagnostics
Calibrator data source Calibrator data are encoded on the 2D barcode of the reagent pack.
o If the reagent pack was produced after the CalSet (calibrator set), the target values
from the reagent pack have priority and are used for generating the calibration
curve.
o If the reagent pack was produced before the CalSet, data followed by target values
from the calibrator card are used for generating the calibration curve.
Control target values Control target values are provided by different data sources.
from different data
sources If a target value for a control for a specific PreciControl lot/reagent lot combination
has been manually, the analyzer uses this value rather than the value read from the
control barcode card or reagent barcode. When determining which control target
value to use, the analyzer applies the following priority rules:
Priority 1: Target values entered manually for a specific reagent lot
Priority 2: Target values read from the reagent barcode
Priority 3: Target vaules read from the control barcode card (or data downloaded
from cobas Link - under development)
If a new reagent lot or control lot is then placed on the analyzer, it uses the control
values encoded in the barcodes for these new lots.
This section describes all reagents necessary to run the cobas e 411 analyzer, and the
reagents that are specific for each available test. The available tests are classified into
different groups:
o Thyroid
o Fertility
o Cardiac
o Oncology
o Infectious disease
o Anemia
o Diabetes
o Bone
o Other
e For more information, see User access levels in the Software description section of the
online Help.
Diluents
For most tests where dilution may be necessary, use Universal Diluent or
MultiAssay as diluent. However, some tests—including Anti-HAV, Estradiol,
Progesterone, and NSE (neuron-specific enolase )—require specific diluents.
Reagents for cobas e 411 analyzer tests
e For information on required diluents and recommended dilution factors, refer to the
package inserts of the particular assay reagents.
System reagents
There are specific calibrators for each test. As for quality controls, there are both
controls covering multiple tests and controls that are specific for only a single test.
e For information on required calibrators and controls, refer to the respective package
insert.
To get information about what calibrator and controls are currently needed for
calibration or QC, print a Calibration/QC Load List from the software.
Reagent packs
The principal reagent container for the cobas e 411 is the cobas e pack.
e For additional information on the packaging and barcoding of reagents, see the Product
labeling section in the System overview chapter of the cobas e 411 analyzer Operator’s
Manual.
Product labeling
A cobas e pack is a reagent pack that consists of three separate, capped reagent
containers. The cobas e 411 can open and close these caps automatically. There is an
individual reagent pack available for each test.
Product labeling
Each reagent pack is equipped with a barcode label. The barcode label contains
reagent, control, and calibration information. The reagent barcode labels are in a
unique format. The symbology uses portable data files (PDF) and is called PDF417.
Traditional linear barcodes serve as a link to relevant information stored in a
database. However a PDF417 is a two-dimensional, stacked barcode encoded to
contain an entire data record. The large amount of data that can be encoded allows all
instrument settings to be included, as well as the master calibration curve and
additional information for the assay. From this master calibration curve and from
the operator 2-point calibration, the analyzer derives the update of the master
calibration curve.
“Every PDF417 symbol (barcode) contains two error detection codewords that are
used like the check digit in linear barcode symbologies to detect decode errors and
verify that all data has been read and decoded accurately. Additionally, PDF417
provides error correction in the event that portions of the symbol have been
damaged, destroyed or are unreadable.” (a)
It is a combination of this error detection and error correction that ensures a
reliable barcode. There should only be a small number of exceptional cases when
barcodes are so badly damaged that the analyzer cannot read them. If the barcode
cannot be read
(a) 1. Itkin S, Martell J. A PDF417 Primer: A Guide to Understanding Second Generation Barcodes and
Portable Data Files. Bohemia, NY: Symbol Technologies, Inc; 1992:17-18.
Data links
and the reagent lot has previously been used by the analyzer, you can manually enter
the 15-digit number found on the reagent barcode label into the software.
Data links
This following table illustrates the information that may be encoded on the barcode
labels. Background colors are used to indicate calibration links that exist between
information on separate barcodes.
The exact information encoded onto the barcode labels is not detailed here, as this
information is propriety to Roche Diagnostics. Items that are not directly linked are
mapped within the cobas e 411 software or user interface.
Data links
e For further information about reagent checklists, see Reagents, calibrators, and controls
in the Troubleshooting chapter of the cobas e 411 analyzer Operator’s Manual.
Calibration
Calibration
The calibration curve produced from the barcoded master calibration and the
measured calibration is specific to each reagent lot and, in some cases, to an
individual reagent pack. The condition of the analyzer and the reagents influence the
system-specific calibration. The result of a calibration is validated automatically by
the analyzer and can be further validated by the operator.
Master calibration
Master calibration
RD Development
Effort
RD Production
Customer
A reference standardization curve utilizing master test kit reagents and certified
reference standard material [for example, World Health Organization (WHO)
reference material] is measured at Roche Diagnostics. This curve uses 10 to 12 points
(n = 10 to 12). The reference standard curve is the basis for the production of
master calibrators.
A lot-specific master calibration curve (n = 5 or 6) is measured at Roche Diagnostics
using lot-specific test kit reagents and master calibrators. The shape of the lot-specific
master curve is characterized by a four-parameter Rodbard function. The data
characterizing this curve are stored in the lot-specific reagent barcode. Lot-specific
calibrator assigned values (CalSet assigned values) are read off the lot-specific master
calibration curve and are encoded in the barcode label of the reagent pack.
At the customer site, the calibration results from two calibrators that were measured
under routine conditions are mathematically combined with the encoded data from
the 2D barcode. From this combination, the system determines a lot calibration or
reagent pack calibration from which the concentration of measured samples is
reliably calculated.
Lot calibration
Lot calibration
A lot calibration (L-Calib) is a calibration performed with a fresh reagent pack that
has not been on the analyzer longer than 24 hours and for which all calibration
validation criteria are acceptable. Reagent-specific calibrators are used to update two
of the four Rodbard curve-defining parameters. This adjusts the curve to match the
original lot-specific calibration curve.
The lot calibration is valid for all other reagent packs of the same lot, provided
these reagent packs were stored as specified in the package insert and have not been
on the analyzer for longer than seven days.
e For further information, see Calibration factor (quantitative assays only) on page C-28.
A reagent pack calibration (R-Calib) is performed with a reagent that has been on
the analyzer for more than 24 hours.
A reagent pack calibration is valid for one specific reagent pack only. The reagent
pack calibration is compared to the most recent stored L-Calib for validation.
Difference between lot and reagent calibration
The following diagram illustrates the process for determining whether a lot or reagent
pack calibration is required:
Reagent pack
Valid for all reagent packs of the same lot Calibration Valid for this reagent pack only
Controls
and samples before operator andwill
release samples after
use the operator
previous lot release will use this reagent pack calibration
calibration Controls and samples
Controls and
after system release will
samples
use this reagent pack
calibration
Figure C-8 Overview of the lot and reagent pack calibration process
Calibration procedures
Calibration procedures
The following table contains examples of the procedures to be followed for various
scenarios when more than one reagent pack is used for one assay:
Calibration stability
Calibration can be performed more frequently if required. This may be because of local
regulations, or before performing specific types of test.
Calibration validation
The calibration of a test can be easily identified on the Calibration > Status screen.
If the test is highlighted in red, this indicates that both of the calibrator measurements
have failed or the calibration factor is outside the range 0.9-1.2 . There is no red
highlighting if the calibration was successful.
Calibration assessment
Calibration assessment
Missing values
Duplicate determinations of two calibrators are used to adjust the master calibration
curve stored on the reagent pack barcode. Therefore, you must have a minimum of
n-1 values for all calibrator replicates measured (n = total number of calibrator
replicates. For any current Elecsys assay, this number totals 4.). Currently, all
Elecsys reagents use only two calibrators. This box can accommodate up to five
calibrators.
Check to see if any alarms occurred during calibration that may have caused the
missing values. Treat any questionable calibration according to laboratory policy.
A curve position check against the most recent lot calibration produces a
calibration factor. This box displays a number that represents this factor.
Each new lot calibration (L-Calib) uses a calibration factor of 1. For all subsequent
reagent pack calibrations (R-Calib), a new calibration factor is calculated. The
calibration factor is the quotient of the slopes of the actual performed calibration and
the related stored calibration.
The calibration factor criterion is used only in validating reagent pack calibration (R-Cal).
The calibration factor is set to 1 at each new lot calibration. The following reagent
pack calibrations are compared with the last measured lot calibration. The calibration
factor is the ratio between the calibrator signals (difference of CalSet 1 and CalSet
2) of the lot and reagent pack calibration. The calibration factor is only used as a
calibration validation criteria and not used for sample calculation.
The following formulae show the relationship between lot calibration and reagent
pack calibration:
t1
Calibration factor for each Lot calibration = --- = 1
t1
t1
Calibration factor for Reagent pack calibration = =
tr actual CalSet 1r signal – actual CalSet 2r signal
CalSet 1l signal – CalSet 2l signal
Example:
r Reagent calibration
l Lot calibration
This simplified formula is only valid when the same calibrator concentrations are used for the
reagent pack and lot calibration. If these calibrator concentrations are different, the calibration
signals of the standardization have to be considered.
Calibration assessment
Minimum signal
The measured signal of the calibrator replicate must be above the minimum value.
Values are test dependent and are encoded in the reagent barcode. Currently, all
Elecsys reagents use only two calibrators. This table can accommodate up to five
calibrators.
Check to see if any alarms occurred during calibration that may have caused a
calibrator replicate to have an unacceptable minimum signal. Treat any questionable
calibration according to your laboratory policy.
The measured signal of the calibrator should fall between the designated minimum
and maximum signals. Minimum and maximum signals are test dependent and are
encoded in the reagent barcode. If all calibrator replicates were sampled with no
errors, this box displays four dashes, representing the calibrator replicates.
Defined as the difference in percent of the values between calibrator 1 and 2. This
difference must amount to at least 30% for the calibration to be accepted.
The difference between the negative and positive calibrator signal values must be
greater than the allowable value limit. This limit is test dependent and is encoded
in the reagent barcode. The minimum acceptable difference is listed as OK or NG
(Not Good).
Deviation of duplication
The deviation of duplicate measurements is a check of the signal values for each
replicate of a calibrator. If the difference between the duplicate measurements is too
great, the appropriate calibrator is flagged. The signal values are used to calculate the
mean value of the duplicate measurements.
System errors
Cal.
The actual measurement signal levels of Cal 1 and Cal 2, used to calculate the mean
value of the duplicate measurements. Two measurements are taken:
1. Signal The actual signal level of the first measurement of Cal 1 or Cal 2. The mean of the first
and second measurements is used in the calculation of the calibration curve.
2. Signal The actual signal level of the second measurement of Cal 1 or Cal 2. The mean of
the first and second measurements is used in the calculation of the calibration
curve.
The target value of the calibrator is encoded in the 2D barcode of the reagent pack.
Qualitative assays are calibrated by a scaling factor, or the cutoff value. The actual
cutoff value is calculated by means of the cutoff formula on the basis of at least one
reactive or nonreactive calibrator. Each sample receives a scaled result value, the
cutoff value, which allows for the classification of samples being reactive or
nonreactive.
The following is a description of the different methods utilized by the cobas e 411
analyzer for calculating results. To calculate quantitative tests, the system uses the
following three calibration functions to convert measured signals into
concentrations:
o Rodbard function
o Linear calibration function
o Linear reciprocal calibration function
The calibration function used by the system is encoded in the 2D barcode on the
appropriate reagent pack. The calculations are performed automatically by the
analyzer, including the correction for samples diluted by the analyzer.
Calibration of quantitative assays
Rodbard function
The relation between the measured signal and the corresponding analyte
concentration is given the following equation:
x Sample concentration
y Signal
Parameters b and c define the shape of the curve and parameters a and d define the
position of the curve.
Under the controlled conditions of automation on the analyzer, the shape of the
calibration curve is very stable and, therefore, it is possible to calibrate this nonlinear
function with only two calibrators and the information of the shape parameters b and
c. The curve position parameters a and d are calculated with each calibration. Such a
calibration is called 2-point calibration.
The following inverse formula is used to determine the sample concentration based
on its signal.
a – y
Equation C-2 x = b 1/c
y – d
y Signal
a,b,c,d Rodbard function parameters
x Sample concentration
Calibration of quantitative assays
The relation between the measured signal and the corresponding analyte
concentration is given the following equation:
Equation C-3 y = bx+a
y Signal
x Concentration
a,b Calibration curve parameters (y-intercept and slope)
Calibrations using a linear calibration curve are always performed using two
calibrators.
The following inverse formula is used to determine the sample concentration based
on its signal.
Equation C-4 y–a
x =
b
x Sample concentration
a,b Calibration curve parameters
y Signal
The relation between the measured signal and the corresponding analyte
concentration is given the following equation:
1
Equation C-5 -- = b x +
ay
y Signal
x Concentration
Calibrations using a linear reciprocal calibration curve are always performed using
two calibrators.
The following inverse formula is used to determine the sample concentration based
on its signal.
Equation C-6 1–ay
x =
by
x Sample concentration
a,b Calibration curve parameters
y Signal
Calibration of
qualitative assays Calibration of qualitative assays
In order to calculate the result of a qualitative assay (cutoff test), the system compares
the effective signal of the measurement Seff with the cutoff signal of the calibration
SCutoff . For that purpose, a cutoff index CutoffIndex is calculated as the ratio between
the effective signal and the cutoff signal as follows.
Seff
Cutoff
Equation C-8 = ----------------
Index S Cutoff
If the effective signal of the measurement Seff equals the cutoff signal of the
calibration SCutoff , the cutoff index CutoffIndex equals 1. For effective signals being
lower or higher than the cutoff signal, the cutoff index is smaller or larger than 1,
respectively.
In order to evaluate the reactivity of a sample, the 2D barcode contains defined limit
values. If the cutoff indices, which were calculated from the effective signals, lie
between the lower limit (LL) and the upper limit (UL), no decision can be made
regarding the reactivity or non-reactivity of the sample (borderline).
The test result is evaluated as follows, depending on the test principle (sandwich tests
show a positive slope, and competitive tests show a negative slope).
Result calculation for qualitative assays
The analyzer automatically calculates the cutoff based on the measurement of Cal1
and Cal2. The results of a sample is given either as reactive, nonreactive, or
borderline, as well as in the form of a cutoff index.
For sandwich assays:
o Samples with a cutoff index > 1.0 are considered to be reactive.
o Samples with a cutoff index < 1.0 are considered to be nonreactive.
For some assays a grey zone is introduced.
For competitive assays:
o Samples with a cutoff index > 1.0 are considered to be nonreactive.
o Samples with a cutoff index < 1.0 are considered to be reactive.
Quality control D
This chap
provides
overview
assignme
control t
values fo
cobas e
analyzer
5 In this
chapte
Chapt
Control t
value (fir
assignme
..............
33
Roche D
quality c
principle
..............
33
Control r
specifica
..............
33
cobas e 411 5 Quality control concept
Table of contents
Roche Diagnostics
COBI-CD · Version 1.0 D-4
Control target value (first) assignment
If the difference between the medians of the target values for cobas e 411 analyzer controls is less
than 1 SD, the mean of these medians is used as the target value.
Case 1 All controls are within the target range: < +/- 1 SD
The target values of the existing control barcode cards are used for this reagent lot,
which means, the control values are not changed when using this reagent lot.
Case 2 The controls are out of the target range: > +/- 1 SD for this reagent lot
The target values of the controls are on the reagent pack barcode, which means the
specific target values for reagent lots exist and the control values on the control card
are not valid. As a consequence, an extra information sheet is put inside the reagent
kit indicating the re-assigned values and the new values are stated on the reagent pack
barcode.
valid.
5 Quality control concept cobas e 411
Control target value (first) assignment
Due to the priority rules with a new reagent lot, the target value of a control will
not be taken from the control barcode card or the reagent barcode if the target value
has been entered manually for this assay.
The main point of each standardization action is to receive the same human
serum recovery independent of the reagent lot. Multi-analyte controls are spiked,
stripped and preserved and unfortunately, do not always react in the same way.
Roche Diagnostics
D-6 COBI-CD · Version 1.0
Index E
cobas e 411 Index
Index
A
Abbreviations, 4 Cleaning
Analyzer cycles – measuring cell, A-9
– automatic, A-21 Competitive principle, C-6
Approvals, instrument, 2 Contact addresses, 3
Aspiration Control target value assignment
– reaction mixture, A-9 – Roche controls, D-5
Assay principles Copyrights, 2
– bridging principle, C-10 Cutoff value, C-30
– competitive principle, C-6
– sandwich principle, C-8 D
Assay sequence, A-8, A-14 Data links
– calibration, C-19
B Deviation of duplicate calibration measurement, C-29
Barcode
– calibration information, C-19 E
Borderline range, C-30 ECL (electrochemiluninescence)
Bridging principle, C-10 – advantages of, B-10
– assay principles, C-5
C – measuring cell, B-9
Calibration – measuring principles, B-5
– barcode information, C-19 – reaction, B-6
– curve, C-21 – signal generation, B-8
– data links, C-19
– factor, C-28 F
– introduction, C-21 Finalization, A-9
– master, C-22 Flow
– procedures, C-25 – operation, A-13
– qualitative assays, C-33 Function
– quality criteria, C-27 – linear calibration, C-32
– quantitative assays, C-30 – linear reciprocal calibration, C-32
– reagent pack, C-23, C-24 – Rodbard, C-31
– signal level, C-30 I
– stability, C-26
Immunology calibration
– validation, C-26 – calibration validation, C-26
Calibration assessment, C-27
– lot calibration, C-23, C-24
Calibration quality criteria
– master calibration, C-22
– borderline, C-30
– procedures, C-25
– calibration factor, C-28
– qualitative assays, C-33
– calibration signal level, C-30
– quality criteria, C-27
– cutoff, C-30
– quantitative assays, C-30
– deviation of duplicate, C-29
– reagent pack calibration, C-23, C-24
– minimum acceptable difference in calibrator
– stability, C-26
signal, C-29 Incubation
– minimum difference, C-29 – first, A-8, A-18
– minimum signal, C-29 – second, A-9, A-19
– minimum/maximum signal, C-29
– third, A-9
– missing values, C-27
Initialization process, A-6
– monotony, C-27 Instrument
– slope, C-27
– approvals, 2
– system errors, C-29
– target, C-30
Roche Diagnostics
COBI-CD · Version 1.0 E-3
Index cobas e 411
Q
Qualitative assays
– calibration, C-33
– result calculation, C-33
R
Reaction mixture
– aspiration, A-9
– measurement, A-9
Roche Diagnostics
E-4 COBI-CD · Version 1.0