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cobas s 201 system

Roche cobas s 201 system User manual

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  • What is a batch in the context of Roche cobas s 201 system?
    How are samples and controls tracked during a batch process?
    What types of external controls are required for each batch?
    What is the purpose of pipetting RMECs before sample pipetting?
    What is a large Primary Pool, and how is it created?
    What is the purpose of 2D Pooling, and how does it work?
    How many 2D Pools are created based on the size of the reactive large Primary Pool?
02/2008, version 1.0 2.1
Pipetting 2
Batch Concept
The cobas s 201 system is designed to process samples in batches. A batch
is a collection of samples and controls that are pipetted, extracted, and
amplified and detected together according to the rules for the associated
test specification.
A batch consists of all of the samples and controls in one SK24 rack.
A batch is tracked from pooling through results review using the SK24
rack ID plus a unique batch ID assigned during pipetting.
Samples and controls in the batch are tracked by associating their scanned
barcode IDs with the unique barcode clips that hold S-tubes (during
pooling and sample preparation) and K-tubes (during amplification and
detection).
Figure 2.1
Batch
SK24 Rack ID
Unique S-tube
Barcode Clip
2.2 02/2008, version 1.0
Roche-Manufactured External Controls (RMECs)
Each batch requires Roche-manufactured external controls (RMECs).
The number of RMECs required is test-specific.
MPX testing encompasses five analytes. Five positive RMECs plus one
negative RMEC must be pipetted for each batch. During pipetting, an
aliquot of the negative control is transferred to the S-tube in position 19 of
each SK24 rack. Then, aliquots from each positive control are transferred
to S-tubes in positions 20 through 24 of each SK24 rack (Figure 2.2).
Pipetting of RMECs always occurs before sample pipetting. This
allows the operator to correct any control pipetting errors before
sample pipetting begins.
RMECs are always placed in the last positions in each SK24 rack so
that the entire testing process, from extraction through
amplification and detection, is monitored by control samples.
User-Defined External Controls (UDECs)
The cobas s 201 system allows up to five user-defined external controls
(UDECs) to be assigned to each test. UDEC requirements, including the
control name, barcode pattern, lot number, expiration date, and position
of the UDECs in the SK24 rack, are specified by the laboratory
administrator.
Once UDECs are assigned for a particular test, the operator can determine
whether to include them in a pipetting run.
If included, UDECs are always pipetted in the first SK24 rack.
UDECs are identified in the Roche PDM Pooling Manager and Roche
PDM Data Manager screens and reports.
Figure 2.2
MPX RMECs in an SK24 Rack
Negative
Positive
Controls
Control
Pipetting
02/2008, version 1.0 2.3
Deep-Well Plates
Identical deep-well plates are used as Library Plates and Intermediate
Plates during pipetting of large Primary Pools (Figure 2.3).
Every deep-well plate has a unique barcode label.
Library Plate
A Library Plate can be prepared during Primary Pooling to save an aliquot
from each donor tube in case Secondary Pooling is required.
Secondary Pooling can be performed directly from the donor tubes
if a Library Plate is not prepared or if a particular well in the
Library Plate is unusable.
Use of a Library Plate is an option that is configured during
installation.
The well position that a sample occupies in the Library Plate is
dependent on the number of samples in the run and the type of
pooling that is being performed.
Intermediate Plate
An Intermediate Plate is required when pipetting large Primary Pools.
The Intermediate Plate holds interim 12-specimen pools that are then
combined to form a large Primary Pool.
Details of pipetting large Primary Pools are described in the
following section.
Figure 2.3
Library Plate and Intermediate Plate
2.4 02/2008, version 1.0
Primary Pools
A large
Primary Pool
is a multi-specimen (
n
=
24
,
48
, or
96
donor samples)
pool that is created for initial sample testing.
Pipetting large Primary Pools is a two-step process consisting of 1) a Plate
Run and 2) a Batch Run. Each is a separate pipetting run.
Plate Run
During a Plate Run, aliquots from groups of donor samples are combined
in Intermediate Plate wells to create 12-
specimen
interim pools.
The number of donor samples that are loaded must be a multiple of the
final pool size (24, 48, or 96). The maximum number of donor samples
that can be pipetted depends upon the pipettor that is used:
•Up to 864 donor samples can be pipetted when the Hamilton
Microlab STAR IVD Pipettor is used
•Up to 384 donor samples can be pipetted when the Hamilton
Microlab STARlet IVD Pipettor is used
Pipetting a Plate Run for a 96-specimen pool is summarized below.
Pipetting a Plate Run for a 24- or a 48-specimen pool is similar.
• 1 mL of each donor sample is aspirated from the first group of
donor sample tubes and dispensed into wells in a Library Plate. The
process is then repeated with 700 μL aliquots from the same donor
sample tubes, resulting in the transfer of 1.7 mL aliquots of donor
samples to corresponding wells of the Library Plate (Figure 2.4).
•135 μL is aspirated from the Library Plate wells and dispensed into
the first column of Intermediate Plate wells (used to store the
interim pools) (Figure 2.4).
If a Library Plate is not prepared, 135 μL aliquots are pipetted
directly into the Intermediate Plate from the donor sample tubes.
Figure 2.4
Pipetting of the First Group of Donor Samples During the Plate Run
(Example of a Plate Run for a 96-Specimen Primary Pool)
Library Plate
Intermediate Plate
32-Position Donor Tube Carrier
1 mL +700 μL
135 μL
Position 1
Position 32
Pipetting
02/2008, version 1.0 2.5
• The process is then repeated with the next group of donor samples.
1.7 mL of each donor sample in the group is transferred to the next
available wells of the Library Plate, and 135 μL is then aspirated
from those Library Plate wells and dispensed into the same column
of Intermediate Plate wells as the first group of donor samples
(Figure 2.5).
• The process continues until all of the donor samples that are
included in the first large Primary Pool have been pipetted and the
wells in the first column of the Intermediate Plate contain aliquots
of twelve donor samples
• The entire process is then repeated to create interim pools for each
additional large Primary Pool, using additional wells in the
Intermediate Plate for the additional interim pools.
At the end of the Plate Run, the Library Plate(s) contain 1.565 mL aliquots
from each of the donor sample tubes, and the Intermediate Plate wells that
are used each contain pooled 135 μL aliquots from twelve donor sample
tubes.
Figure 2.5
Pipetting of the Next Group of Donor Samples During the Plate Run
(Example of a Plate Run for a 96-Specimen Primary Pool)
Library Plate
Intermediate Plate
32-Position Donor Tube Carrier
1 mL +700 μL
135 μL
Position 1
Position 32
2.6 02/2008, version 1.0
Batch Run
During a Batch Run, aliquots are pipetted from Intermediate Plate wells
into S-tubes to create the large Primary Pool (Figure 2.6):
•500 μL aliquots from two wells are pipetted into one S-tube for a
Primary Pool size of 24
•250 μL aliquots from four wells are pipetted into one S-tube for a
Primary Pool size of 48
•125 μL aliquots from eight wells are pipetted into one S-tube for a
Primary Pool size of 96
Intermediate Plates from more than one Plate Run can be loaded. The
maximum number of Intermediate Plates that can be pipetted depends
upon the pipettor that is used:
•Up to 5 Intermediate Plates can be pipetted when the Hamilton
Microlab STAR IVD Pipettor is used
•Up to 4 Intermediate Plates can be pipetted when the Hamilton
Microlab STARlet IVD Pipettor is used
Figure 2.6
Pipetting of Two 96-Specimen Pools During a Batch Run
Intermediate Plate
125 μL
Pipetting
02/2008, version 1.0 2.7
Repeat Batch Run
A Repeat Batch Run
creates another large Primary Pool to replace one with
an
invalid test result
.
The Repeat Batch Run uses the Intermediate Plate
that was created during the first step (the Plate Run) of the original
Primary Pooling run.
A Repeat Batch Run must be pipetted from the Intermediate Plate.
If an Intermediate Plate well is not available the samples are
scheduled for Resolution Pooling.
Load only one Intermediate Plate to preform a Repeat Batch Run.
During a Repeat Batch Run, aliquots from the same Intermediate Plate
wells used to create the original large Primary Pool (Figure 2.6) are
pipetted into S-tubes to create the replacement Primary Pool (Figure 2.7).
Figure 2.7
Repeat Batch Run for a 48-Specimen Primary Pool
Intermediate Plate
250 μL
2.8 02/2008, version 1.0
2D Pooling
2D Pooling (two dimensional pooling) is used to retest samples from
reactive large Primary Pools. During 2D Pooling, each sample is pipetted
into two different pools with a sample distribution (Figure 2.8) that allows
non-reactive sample(s) to be identified after a single run.
Samples from one Primary Pool can be processed during a 2D
Pooling run.
The number of 2D Pools that are created depends on the size of the
reactive large Primary Pool:
Figure 2.8
Sample Distribution in 2D Pools
(Example of 2D Pooling for a 48-Specimen Primary Pool)
Library Plate
Primary Pool 2
Primary Pool 1
Six-Specimen Pools
S-tube Positions 1-8
Eight-Specimen Pools
S-tube Positions 9-14
Pool Size Number of 2D Pools Aliquot of each Sample
24 4 pools of 6 specimens
6 pools of 4 specimens
167 μL
250 μL
48 8 pools of 6 specimens
6 pools of 8 specimens
167 μL
125 μL
96 Two batches each containing:
8 pools of 6 specimens
6 pools of 8 specimens
167 μL
125 μL
Pipetting
02/2008, version 1.0 2.9
Pipetting a 2D Pooling run for a 48-specimen pool is summarized below.
Pipetting a 2D Pooling run for a 24- or a 96-specimen pool is similar.
If a Library Plate is not used, if a Library Plate well is invalid, or if a
Library Plate well contains insufficient volume, aliquots can be
pipetted from donor sample tubes.
•167 μL aliquots of donor samples are aspirated from four wells in
the first column of the Library Plate and dispensed into four
S-tubes, starting in position 1 of the SK24 rack (Figure 2.9).
•125 μL aliquots of donor sample are then aspirated from the same
four Library Plate wells and dispensed into a single S-tube in
position 9 of the SK24 rack.
•167 μL aliquots of donor samples are aspirated from four wells in
the next column of the Library Plate and dispensed into four
S-tubes, starting in position 5 of the SK24 rack.
•125 μL aliquots of donor sample are aspirated from the same four
Library Plate wells and dispensed into the single S-tube in position
9 of the SK24 rack.
At this point, the S-tubes in positions 1 through 8 of the SK24 rack
each contain an aliquot of a single donor sample, and the S-tube in
position 9 contains an aliquot of eight donor samples (Figure 2.9).
Figure 2.9
Pipetting the First Eight Wells
(Example of 2D Pooling for a 48-Specimen Primary Pool)
Library Plate
Position 9
167 μL
125 μL
2.10 02/2008, version 1.0
• The process is then repeated for the next eight wells on the Library
Plate. 167 μL aliquots of donor samples are again dispensed into
the S-tubes in positions 1 through 8 of the SK24 rack, and 125 μL
aliquots are dispensed into a single S-tube in position 10 of the
SK24 rack (Figure 2.10).
• The process continues until all donor samples have been pipetted.
When the run is completed, the S-tubes in positions 1 through 8 of
the SK24 rack each contain aliquots of a six donor samples, and the
S-tubes in positions 9 through 14 each contain aliquots of eight
donor samples.
Figure 2.10
Pipetting the Next Eight Wells
(Example of 2D Pooling for a 48-Specimen Primary Pool)
Library Plate
SK24 Rack
167 μL
125 μL
Pipetting
02/2008, version 1.0 2.11
Confirmation Pooling
Confirmation Pooling is used to retest samples from reactive large
Primary Pools when there is additional evidence (e.g., a positive serology
test) that suggests one or more samples in the Primary Pool is reactive.
Confirmation Pooling is also used to retest Short Pools.
Samples that are suspected to be reactive are pipetted in individual pools.
The remaining samples are pooled in multi-specimen pools (Figure 2.11).
Samples from one Primary Pool can be processed during a
Confirmation Pooling run.
Up to four samples from a Primary Pool of 48 or 96 can be selected
for pipetting in single-specimen pools. Up to two samples from a
Primary Pool of 24 can be selected for pipetting in single-specimen
pools.
The multi-specimen pools are prepared as 12-, 11-, 6-, or 4-specimen
pools, depending on: 1) the number of samples in the Primary Pool and 2)
the number of donor samples selected for individual testing. The volume
of sample that is aspirated for each size multi-specimen pool is
summarized below:
Figure 2.11
Sample Distribution in Confirmation Pools
(Example of Confirmation Pooling for a 48-Specimen Primary Pool)
Library Plate
Primary Pool 2
Primary Pool 1
Multi-Specimen Pools
S-tube Positions 1-4
Single-Specimen Pools
S-tube Positions 5-8
Multi-Specimen Pool Size Aliquot of each Sample
12-specimen 92 μL
11-specimen 92 μL
6-specimen 167 μL
4-specimen 250 μL
2.12 02/2008, version 1.0
Pipetting during Confirmation Pooling is summarized below:
If a Library Plate is not used, if a Library Plate well is invalid, or if a
Library Plate well contains insufficient volume, aliquots can be
pipetted from donor sample tubes.
• Aliquots (92 μL, 167 μL, or 250 μL) of donor samples are aspirated
from wells in the first group of presumed non-reactive Library
Plate wells. Each sample aspiration is dispensed into an S-tube,
starting in position 1 of an SK24 rack.
• The process is repeated, skipping the wells (or donor tubes) for any
donor samples that are suspected of being reactive (Figure 2.12).
Figure 2.12
Pipetting Presumed Non-Reactive Confirmation Pools
(Example of Confirmation Pooling for a 48-Specimen Primary Pool)
Library Plate
SK24 Rack
92 μL
Pipetting
02/2008, version 1.0 2.13
• After all aliquots of presumed non-reactive donor samples have
been pipetted, 1 mL aliquots of the presumed reactive donor
samples are aspirated from their wells (or donor tubes) and
dispensed into individual S-tubes (Figure 2.13).
Figure 2.13
Pipetting Presumed Reactive Individual Pools
(Example of Confirmation Pooling for a 48-Specimen Primary Pool)
Library Plate
SK24 Rack
1 mL
2.14 02/2008, version 1.0
Resolution Pooling
A Resolution Pool is a single-specimen pool that is created for samples
that 1) have pipetting errors during the Primary Pooling run or 2) are
contained in a reactive Confirmation Pool or reactive 2D Pool. A
Resolution Pool is prepared by pipetting an aliquot from a Library plate
well into its own S-tube.
The laboratory administrator can also elect to use Resolution
Pooling to resolve invalid results.
The sample can be aspirated from the donor tube if a Library Plate
is not available or if the Library Plate well for that donor sample is
unusable.
Up to 36 samples can be processed during a Resolution Pooling
run.
During Resolution Pooling, 1 mL of a sample is aspirated from its well in
the Library Plate and dispensed into a single S-tube.
Figure 2.14
Pipetting Resolution Pools
Library Plate
SK24 Rack
1 mL
Wells selected for Resolution Pooling
/