2. WHY FULLY AUTOMATION IN
BLOOD BANK ?
To achieve “safe & timely blood transfusion”
Faster TAT =>Faster blood transfusion
Reduce labour intensive manual test procedure
Test results directly upload to LIS (reduce
transcription error)
Thus, better quality of blood transfusion
2
3. Personnel shortages, TAT requirements and greater
regulatory demands have provided incentive for blood
services to seek automation
Automated equipments provide the level of quality
assurance required by new regulatory standards
Barcode reduces identification errors by providing
accurate patient identification
3
4. Automated systems from different manufactures
use advanced technologies like CAT (Column
Agglutination Technology) and SPRCA (Solid
Phase Red Cell Adherence)
Popular names for CAT and SPRCA are Gel and
Solid Phase technology respectively
Within the last several years automated methods
for typing blood, screening blood antibodies have
been developed.
4
5. The automated methods have led to a
remarkable improvement in blood safety
The application of automation is now growing
in hospitals and transfusion services, because
human errors can lead to life threatening
consequences for the patients
6
6. Two basic Automation methodologies in blood
bank
I)GEL TECHNOLOGY (column agglutination using
a gel matrix)
II)SOLID PHASE TECHNOLOGY (solid-phase red
cell adherence)
8
7. The equipments that have been fully automated
and currently available in market are
Galileo (Immucor, Inc)- uses SPRCA
Techno (Diamed)- uses CAT
Auto Vue (Ortho Clinic Diagnostics Ltd)- uses CAT (beads)
9
8. GEL TECHNOLOGY
Gel technology is currently approved for ABO forward
and reverse grouping, Rh typing, DAT, antibody screen,
antibody identification and compatibility testing.
HISTORY:
In 1985,the gel test was developed by Dr Yves
Lappierre of France
Media used- Gelatin, acrylamide gel, glass beads
Gel appeared to be ideal material for trapping
agglutinates
Compared with traditional tube technology, the gel
test provides a more stable endpoint.
11
9. PRINCIPLE:
Performed in a specially designed microtube.
Based on controlled centrifugation of RBC through
a dextran-acrylamide gel that contains predisposed
reagents
12
17. Measured volumes of serum or plasma or RBCs
are dispensed into the reaction chamber of the
microtube
The card is incubated and then centrifuged.
The reaction chamber is actually a miniature test
tube, providing an area for the sensitization of
RBCs during incubation.
The shape and length of the column provides a
large surface area for prolonged contact of the
RBCs with the gel particles during centrifugation.
20
18. The gel particles are beads of dextran-acrylamide
that make up 75% of the gel-liquid mixture that is
preloaded into each microtube.
The gel particles are porous, they serve as a
reaction medium, filter, sieving the RBC
agglutinates according to size during
centrifugation.
Large agglutinates are trapped at the top of the gel
and are not allowed to travel through the gel during
the centrifugation of the card .
21
19. Agglutinated RBCs remain fixed or suspended in
the gel, while unagglutinated RBCs travel
unimpeded through the length of the microtube,
forming a pellet at the bottom following
centrifugation.
The gel test reactions are stable, allowing
observation for up to 3 days.
22
23. Negative reactions may appear mixed field when
incompletely clotted serum samples are used in
the gel test
Fibrin strands may trap unagglutinated RBCs
Before interpreting mixed-field, clinical history
should be considered
Ex- recently transfused or bone marrow transplant
recipients are expected to have mixed populations of
RBCs and commonly produce this
28
24. TESTS APPROVED BY FDA
Gel technology is currently approved for
ABO forward and reverse grouping
Rh typing
DAT
Antibody screen
Antibody identification
Compatibility testing
29
25. The ABO blood grouping card contains
gels that include anti-A, anti-B, and anti-A,B for
forward grouping.
Microtubes with buffered gel are used for ABO
reverse grouping.
The Rh typing card uses microtubes filled with gel
containing anti-D.
Microtubes filled with gel containing anti-IgG are
used for compatibility testing, antibody detection,
and identification.
30
26. ADVANTAGES
Standardization is one of the major advantages,
(there is no tube shaking to resuspend the RBC
button)
It includes simple standardized procedures, no
wash steps.
Decreased sample volume needed for testing.
Provides stable, well defined end points of the
agglutination reaction
More objective, consistent and reproducible
interpretation of the test results
32
27. DISADVANTAGES
Need to purchase special incubators and
centrifuges to accommodate the microtube cards
used for testing.
A specific pipette must be used to dispense 25 mL
of plasma or serum and 50 micro L of a 0.8 percent
suspension of RBCs into the reaction chambers of
the microtubes.
Ideally suited to individuals who have been cross
trained to work in the blood bank
33
29. In 1978 Rosenfield and coworkers were the first to apply
the principle of solid-phase immunoassay to RBC typing
and antibody screening tests.
SPRCA (solid phase red cell capture assay) was
developed commercially for the detection of RBC and
platelet related antibodies.
In these test systems,one of the test reactants (either
antigen or antibody) is bound to a solid support (usually a
microtiter well) before the test is started.
The ability of plastics, such as polystyrene to absorb
proteins from solutions and bind them irreversibly made
SPRCA possible.
35
HISTORY
30. SPRCA was developed by trade name Capture®
by Immucor
In the Capture® immucor technology, tests were
adapted to microplate wells, either as full 96-well U-
bottomed plates or as 1x8 or 2x8 strips of U-
bottomed wells
Capture-R® for the detection of RBC antibodies and
Capture-P® for the detection of platelet antibodies.
To perform these tests, a laboratory centrifuge
capable of holding 96-well microplates is required,
along with a microplate incubator and an
illuminated reading surface or microplate reader
36
31. PRINCIPLE
Based on SPRCA
The tests uses chemically modified microplate test
wells in which intact reagent RBCs are bound to the
microwells before starting the test.
Patient serum or plasma and low ionic strength saline
are added to the RBC-coated microwells and
incubated at 370c to allow time possible for antibodies
to attach.
After incubation, the wells are washed with pH-
buffered isotonic saline to remove unbound proteins
37
32. Wells are added with an indicator of anti-IgG–
coated RBCs, and the microwells are centrifuged.
Indicator cells are AHG coated RBCs
Centrifugation forces the indicator RBCs to
contact the immobilized sensitized reagent RBCs.
Positive test show adherence of indicator RBCs to
part or the entire well bottom, depending on the
strength of the reaction
38
33. In second generation tests, RBC membranes are
bound to the microplate test wells and air dried
Capture R® immucor is first generation solid
phase test, Capture R® Ready-Screen/Capture-
R® Ready-ID® are second generation tests
Solidscreen® is a solid phase test that detects
antibody using microplate wells that are coated
with polyspecific antihuman globulin
39
34. TEST REACTIONS
If antibody has attached to antigen , the indicator
cells will form a monolayer of RBCs
No antibody, nothing attached to antigen and
indicator cells will form a clearly delineated button
at the center of microplate well
Positive tests show adherence of indicator RBCs
to part or all of the well bottom, depending on the
strength of the reaction
40
37. ADVANTAGES
Standardization is the major advantage of solid-
phase technology.
Solid-phase technology provides stable, well-
defined endpoints of the reaction.
Ease of use
No predilution of reagents is required.
It is possible to test hemolyzed, lipemic, or icteric
samples.
The enhanced sensitivity makes the detection of
weak alloantibodies easier.
43
38. DISADVANTAGES
The major disadvantage is
the need for a centrifuge that can spin microplates,
a 37°C incubator for microplates
a light source for reading the final results.
Increased sensitivity may also be a disadvantage
in as much as solid phase may detect weak
autoantibodies that other systems miss
44
39. Solid phase technology for
immunohematology
Capture has proved to be most dependable technology as it has
following features:
ACCURACY:
IgG specific technology- detects clinically significant antibodies
Sensitivity and specificity are higher
Automated washing of hemolyzed, icteric, lipemic samples
assures validity
Controls with each run assures validity
45
40. EASE OF USE:
Antigen are pre coated on test wells
No pre-dilution of samples or reagents
Same method and procedure for all assays
46
41. COST EFFICIENCY:
Higher lab productivity
Longer shelf life of coated antigens
Cost benefit of single antibody screen of patients
vs. number of crossmatches
Competitively priced
Easily automated or upgraded
47
42. PARAMETER ‘CAPTURE’ SPRCA OTHER TECHNOLOGIES
SENSITIVITY Highly for IgG antibodies Less
SPECIFICITY More False positivity is higher
SHELF LIFE 3-4 months 4-5 weeks
STANDARDIZATION Standardized Not
APPROVALS US FDA approved Non- US FDA approved
PLATELET
ANTIBODY
SCREENING AND
CROSSMATCH
available Not available
48
43. CURRENT TESTS AVAILABLE
Test Gel test Solid Phase
ABO- Forward Yes No
ABO- Reverse Yes No
Rh typing Yes No
Antibody screen Yes Yes
Crossmatch Yes Yes
Antibody identification Yes Yes
49
45. Reaction
readings
Quantitative:
1 to 4
Quantitative:
1 to 4
Semiquantitative
g: strong
positive,
positive,
Negative, no MF
Stable reactions No yes (2–3 days) Yes (2 days)
Quality control Positive and
negative control
Lot number of
cards and diluent
on day of use
LISS color
change, positive
and negative
control
Special
equipment
No Yes Yes
Automation No Yes Yes
51
46. SENSITIVITY
Technologies has been shown to detect at least as
many IgG Ab as conventional test tube methods
Sensitivity can be modified by pretreating the test
cells or monolayer with enzymes
Increased sensitivity is advantage when low titered
clinically significant antibody present
52
47. QUALITY CONTROL
GEL TEST
Uses special dispensers to prepare the RBC
suspensions and special pipettes to add measured
volume of plasma/serum
Each lot number of cards and diluent should be
tested on the day of use to confirm that the test
cards and diluted reagent RBC’s are reacting as
expected
53
48. SOLID-PHASE TECHNOLOGY
Recommends including a positive and a negative
control with each batch of tests
LISS formulated to detect the addition of plasma
by a color change from purple to blue when
plasma is added
54
49. AUTOMATION
Automated instruments use Gel and solid phase
technologies to increase the efficiency and
productivity of donor testing
They include ABS2000, ROSYS Plato(SPRCA),
Galileo,Diamed and ProVue by ortho clinical
diagnostics
These systems are capable of performing ABO/Rh
and antibody screens in the processing of donor
blood
55
51. The ABS2000 is credited as being the first fully
automated walk away system
It performs ABO/Rh using hemagglutination and
antibody screens/ crossmatches using solid phase
technology
The ABS2000 uses a bardode scanner to log
reagents and samples, transfer specimens by
automated pipetting, prepare RBC suspensions,
incubate, wash, centrifuge, read and interpret
results
Performs low to medium volume workloads
57
54. Galileo is a fully automated walk away system
which uses solid phase microplate technology to
perform all tests.
It uses SPARC technology for all coombs based
testing like
AHG cross matching
antibody screening
antibody identification
DAT
For routine blood grouping testing, it uses
hemaglutination technology
61
55. Special and unique feature is availability and
automation of Platelet Antibody screening and
Platelet Cross matching
Manufactured by Immucor Inc, USA and used in
many countries
62
56. It has different components like
Centrifuge
Incubator
Washer
Camera
Sample and reagent loading bay
Microplate loading tower
which all are coordinated with each other to process
testing and no manual intervention is required
It has a sample capacity of 224 samples and
number of reagents can be loaded depending on
nature of tests performed
63
57. All reagents are bar-coded which helps in easy
identification of the same by instrument itself
There are various incubators of different
temperature available to carry different nature of
testing like blood grouping and antibody screening
simultaneously
Thus useful in institutions where different
parameters are performed simultaneously
Test results with images of microplates and all
other relevant details are continuously stored in
memory of instrument which can be reviewed as
and when required
64
58. Advantages
High loadging capacity- efficient to carry out heavy
workload
The capture technology used in Galileo is IgG
specific and hence provides high specificity
The capture technology used in Galileo is highly
sensitive
It has option to perform extended phenotyping for
all clinically significant antigens like Rh,Duffy,MNS
etc.
65
59. Automation of Platelet Antibody screening and
Platelet Cross matching makes it unique
The high processing speed of Galileo helps in high
through put of large batch testing within short time
The cell panels used for antibody screening and
identification in Galileo are in dried form coated on
microplate which increases the shelf life of these
panels for around 10-12 weeks
The washing step involved in assay procedure
enables the use of lipemic, icteric and hemolysed
samples
66
60. The system liquid used is 0.9% normal saline and
it can be replenished even during the test run
It has an option of online remote access through
internet for technical support and it can be used for
diagnostic purpose
Test results can be reviewed in four different
patterns by plate images, by grading of reactions,
by reaction strength only and by straight result as
positive and negative
The inbuilt quality control run as positive and
negative controls ensures validity of test results
67
61. Disadvantages
Closed system- no other reagents can be used other
than those by manufactures
If test run not managed properly, can lead to wastage
of consumables
Cost effectiveness must be considered as the
consumables are usually priced more
Internet connection is required for remote technical
support
68
62. The indicator cells used in coombs testing has
onboard shelf life of 24 hours
Archiving process must be done after regular
interval of 10-15 days to store all test results
69
65. ADVANTAGES
Has two working stations which function
independently and can be useful if one station is
not working
Gel technique is used is more sensitive
Interpretation of test results are done automatically
and image cards are available to review the results
The STAT position is available for urgent sample
processing
72
66. DISAVANTAGES
Not true continuous access and no sample can be
loaded during centrifugation of cards
Sample lodging capacity is of 36 samples
The blood grouping card used does not include
Anti-Ds from two different sources as
recommended by NACO, NBTC, BCSH and AABB
neither it has O cells in reverse grouping
The blood grouping card used also does not
include Anti-A,B which is essential for weak
subgroup of A group
73
67. Instrument lacks the criteria of automated validation
of test results as proper negative and positive
controls are not available during test run
Cards have to be manually placed- ? Fully automated
It is not possible to determine the presence of
clinically significant IgG antibodies when they are
present with cold antibodies as they mask the
presence of IgG reactivity in Gel Cards
The blood grouping profiles are fixed in gel format
and cannot be modified or changed according to
different needs of different blood banks
74
70. Uses Bead cards
ProVue is a walk-away instrument with capacity for
48 samples and 16 reagents
Safety features includes bar-code tracking system,
3 camera that record sample, reagent and card
identification
A camera in the instrument performs image
analysis and uses a mathematical algorithm to
interpret the results
77
71. ADVANTAGES
STAT Position is available for urgent testing of any
sample
Bead cards are sensitive
Partially continuous random access makes it better
in handling nature of testing simultaneously
Few reagents used
78
72. DISADVANTAGES
The Weak D testing not available
Loading capacity- 36
All samples scanned manually by hand barcode
scanner to feed samples in instrument
All samples centrifuged manually prior to loading
samples in instrument
79
73. The sample must be at least 1 ml for processing in
machine which is practically not possible in cross
matching for donor samples from tube segments
If any error occurs like waste container full, the
instrument stops working and waits for technician
to take required action
The machine produces many equivocal reactions
which must be edited manually by operator by
visual interpretation of cards seen on screen
80
74. AUTOMATED IDENTIFICATION
METHODS
BARCODES
A barcode comprises a series of vertical bars and
spaces arranged in various combinations to
represent different characters.
By combining the numbers, letters and other
characters, a series of barcodes can be built up to
represent donation numbers, blood groups and
various blood products
Barcodes have been very good for the blood bank
community. They offer high speed and accurate
data collection in a life-or-death application.
82
75. An eye readable number or description is included
with the machine- readable code.
Device which will interpret barcodes pass a beam
of light across the code making use of 2 levels of
optical reflectance viz. The black bars and white
spaces
83
76. ELISA
First test developed for the detection of
HIV antibodies in 1985 and currently widely
used test for serodiagnosis of HIV infection
Three types
Indirect ELISA
Competitive ELISA
Sandwich ELISA
Based on sandwich principle involving the use
of solid phase
84
77. Test sample is added to the microwell and
incubated
At the end of incubation period, the unbound
antibody/ antigen is removed by washing and
conjugate is added and incubated
Ag if present in the sample allows enzyme
conjugate to bind to the solid phase and build a
sandwich of Ab-Ag-Ab
Excess conjugate is washed away and chromogen
substrate is added for enzyme action to produce
color
85
78. The intensity of color is directly propotional to the
enzyme activity which in turn is propotional to the
Ag concentration in the sample or control
Result can be visualised or by ELISA reader
To determine reactive or non reactive important to
calculate the cut off value given by manufacturer
86