CURRENT ASPECTS

IN BLOOD BANKING

DR. MADHUVAN GUPTA

 ROUTINE WORKING IN BLOOD BANK

1. Blood grouping & irregular antibody detection.

2. Cross matching

3. Blood component preparation and issue

4. Screening of blood for TTI

 BLOOD GROUP

• Blood group is based on presence of specific antigens on the red

cell membrane which are minute glycoproteins and glycolipids of
sufficient molecular weight to act as antigens.

• Red cell antigen can also be present on leucocytes, platelets, body

fluids, saliva, seminal fluid, plasma & most body tissue.
 ABO BLOOD GROUPING SYSTEM

• According to the ABO blood typing system there are

four different kinds of blood types: A, B, AB or O
• Anti AB antibodies are naturally occurring.
 DIFFERENT BLOOD GROUP SYSTEMS
S. No. Blood Group Abbreviation S. No. Blood Group Abbreviation
1 ABO ABO 14 Dombrock DO
2 MNS MNS 15 Colton CO
3 P P 16 Landsteiner Wiener LW
4 Rh RH 17 Chido/Rodgers CH/RG
5 Lutheran LU 18 Hh H
6 Kell KEL 19 Kx XK
7 Lewis LE 20 Gerbich GE
8 Duffy FY 21 Cromer CROM
9 Kidd JK 22 Knops KN
10 Diego DI 23 Indian IN
11 Cartwright YT 24 Ok OK
12 XG XG 25 Raph RAPH
13 Scianna SC 26 JMH JMH
BLOOD GROUP ABO SYSTEM
 RHESUS (RH) SYSTEM

• Rh antigens are transmembrane proteins with loops exposed at

the surface of red blood cells.

• They are named for the rhesus monkey in which they were first
discovered.

• RBCs that are "Rh positive" express the antigen designated D.

• 85% of the population is RhD positive, the other 15% of the

population is running around with RhD negative blood
• Rh antibodies are found only after sensitization

• A person with Rh blood can develop Rh antibodies in

the blood plasma if he or she receives blood from a
person with Rh+ blood, whose Rh antigens can
trigger the production of Rh antibodies.
• Causes of Rh antibody production

• Transplacental haemorrage in Rh negative women carrying Rh

positive fetus due to

• Delivery

• Amniocentasis

• Abortion

• Trauma

• Transfusion of Rh incompatible blood or blood products

 BLOOD GROUPING PROCEDURES

• Slide or Tile method

• Tube method

• I.D. Micro typing system

• Microplate method

• Automation or semi-automatic instrumentation.

POOLED CELL SUSPENSION
 FORWARD TYPING AND REVERSE
TYPING

• Forward grouping / Cell grouping - the unknown test cells are

tested against Anti-A and Anti-B.

• Reverse grouping / Serum grouping - the unknown serum is

tested against known group A and group B cells.

• The results of both must match to confirm the true ABO type of
an individual.
Advantage:

• For emergency purposes

Disadvantage:

• Insensitive for weak antigens in forward grouping and low titre

antibodies in reverse typing.

• Drying effect
 TUBE METHOD

IMMEDIATE SPIN SALINE ROOM TEMPERATURE

In Reverse grouping or serum grouping

In this similar technique is used, to test patient’s serum with

pooled cell suspension of group A cell, B cell and O cells.

Advantage :-

• Allows for longer incubation without any drying.

• Better for ABO grouping than slide technique.

 GRADING OF ABO GROUPING

• Negative: uniform suspension of red

cells.
• Grade 1 (1+): small clumps of red cells
• Grade 2 (2+): large clumps with many
red cells
• Grade 3 (3+): three or four individual
clumps with few red cells
• Grade 4 (4+): one solid clump of red
cells with no free red cells
 I.D. MICROTYPING SYSTEM
Principle:

•Microtubes in the form of cards , filled with buffered dextran gel

are used which may be neutral or impregnated with antisera.

•With negative reaction the red cells pass through the gel upon
centrifugation.

•In positive reaction the agglutinated red cells are trapped on top
of gel or suspended within it.
 MICROPLATE

• It is ideal for testing large number of blood samples.

• This is rapidly replacing test tubes in many

laboratories.

• Microplates are polystyrene plates having 96 small

wells which can hold 200-300 microlitres of reagent.

• Wells are U-type, V- type or flat bottomed.

• Microplates are intended to be disposable however they can be
reused after cleaning them properly making sure that all foreign
proteins are removed.

• Microplates can be incubated & centrifuged.

• More sensitive to detect weaker antigen-antibody reactions.

• Microplates can be adapted for automation.

MICROPLATE
I.D. GEL CARD
Negative reaction
• Red cells pass through gel upon centrifugation

Positive reaction
• Agglutinated red cells trapped on top of the gel.
GRADING
 ADVANTAGES

• Easy to use system.

• Simple method.

• Microtubes are combined on one test card which facilitates

and simplifies labelling.

• Errors are avoided and safety is increased.

• Clean work place. Special racks for sample tubes and typing
cards.
• Versatile system: this card for reverse typing is a very good
example. It contain 3 tubes with neutral gel and 3 tubes with
coombs gel.

• Result can be easily documented by making photocopies.

• Reaction patterns are stable for hours or even for days if cards
are closed with tape & kept in a refrigerator.

• Typing cards allow automation.

• Clear cut results.

• Available for performing blood group serology, antibody

screening (by DAT) and pre transfusion cross match.
Negative reaction:
• Red cells trail from the center of the well.

Positive reaction:
• Cells remain in center or fall as discrete button.
 GRADING

• Negative No agglutination
• Grade 1+ Fine granular appearance visually, but definite
small clumps per low power field
• Grade 2+ Many fairly large clumps with many free cells
• Grade 3+ Majority of cells agglutinated with some free
cells
• Grade 4+ Complete agglutination of all cells
Advantage :

• Ideal for testing large number of blood samples

• Saves time

• Saves cost
CROSS MATCHING
 CROSS MATCHING

Purpose: It is carried out to ensure that there are no antibodies

present in patient’s serum that will react with donor cells when
transfused.
• Cross match is only part of the compatibility test.

• The compatibility test fully consists of:-

- Review of patient’s past blood bank history & records.

- ABO & Rh typing of recipient & donor.

- Antibody screening of recipient’s & donor’s serum.

- Cross match.
Main 2 functions of cross match test are:-

• It is final check of ABO compatibility between donor and patient.

• It may detect the presence of an antibody in patient’s serum

which will react with antigens on donor cells which was not
detected in antibody screening because absence of
corresponding antigen in the screening cells.
• Major cross match: testing of donor red cells with recipient’s
(patient’s) serum, to detect antibodies in patient.

• Minor cross match: testing patient’s red cells with donor’s

plasma to detect antibodies in donor plasma.
MAJOR CROSS MATCH TECHNIQUES

• Immediate spin technique

• Saline room temperature technique

• Albumin addition technique at 37 ˚ C

• Indirect Antiglobulin technique

• Albumin & LISS can be used with AHG to increase sensitivity.
COOMB’S TEST
 PRINCIPLE OF ANTIGLOBULIN TEST

• The incomplete antibodies (IgG) attach to red cell membrane

by the Fab portion of the immunoglobulin molecule (IgG).
• The IgG molecules attached to the red cells are unable to
bridge the gap between sensitized red cells which are
separated from each other by the negative charge on their
surface and the sensitized red cells do not agglutinate.
ADDING OF ANTIGLOBULIN SERRUM COMPLETES THE REACTION
• When AHG serum is added to the
washed sensitized cells, the Fab
portion of the AHG molecule (anti-IgG)
reacts with the Fc portions of 2
adjacent IgG molecules attached to red
cells thereby bridge the gap between
sensitized red cells and cause
agglutination.
 INDIRECT COOMBS TEST
(INDIRECT ANTIGLOBULIN TEST)

• This test is performed to detect presence of Rh-antibodies or other

antibodies in patient’s serum in case of the following:
• Transfusion of Rh positive blood
• Pregnancy, if infant is Rh positive (if father is Rh-positive)
• Abortion of Rh-positive fetus.
 TEST INTERPRETATION

TEST TUBE OBSERVATION CONCLUSION

Positive Control (PC) Agglutination Correctly performed
test procedure
No agglutination Coombs serum may
not be proper.
Repeat the test
Negative Control It should show no agglutination since saline
(NC) does not contain Anti-D or any other
antibodies
Test Serum (T) Agglutination (and if Patient’s serum
PC results are contains anti-D
correct)
FACTORS AFFECTING THE SENSITIVITY OF
IAT

• TEMPERATURE
• SERUM: CELL RATIO
• INCUBATION TIME
• SUSPENDING MEDIUM
 LOW IONIC STRENGTH
INDIRECT ANTIGLOBULIN TEST(LISS/IAT)

• Principle: Use of LISS in IAT increases the rate and degree of

antibody uptake by red cells and reduces the incubation time
considerably.
 DIRECT COOMBS TEST
(DIRECT ANTIGLOBULIN TEST)

• This test is performed to detect anti-D antibody or other

antibodies attached to the red cell surface within the blood
stream.
• This occurs in the following circumstances:

• Transfusion reactions

• Drug induced red cells sensitization

• Autoimmune hemolytic anemia.

DIRECT ANTIGLOBULIN TEST (DAT)

Cell coated in vivo

Washed to remove
unbound globulins.

Addition of anti-human
globulin (AHG) promotes
agglutination after
centrifugation
• The direct antiglobulin test (DAT) detects sensitized red cells
with IgG and complement components C3b and C3d in vivo.
• In vivo coating of red cells with IgG and complement may
occur in any immune mechanism is attacking the patient’s
own RBC’s.
• This mechanism could be autoimmunity, alloimmunity or a
drug-induced immune-mediated mechanism.
 DAT BY GEL METHOD

• Prepare 0.8% Red cell suspension in LISS

• Take 50ul in a microtube of the LISS/ Coombs ID card

• Centrifuged for 10 min

• Examine for agglutination and hemolysis

• Positive reactions were graded from 1+ to 4+

 FALSE POSITIVE RESULTS: DAT & IAT
• In specimens containing potent cold-reactive antibodies
agglutination may occur before adding the AHG reagent.
• Dirty glassware may cause clumping of cells.
• Over centrifugation
• A positive DAT from a clotted sample should be repeated on
an EDTA sample.
• Sample collected from infusion lines may have complement
present on the cells.
ANTIBODY SCREENING

(ANTIBODY DETECTION

& IDENTIFICATION)
• Purpose of the antibody screen - to detect
red blood cell antibodies other than anti-A or
anti-B.

• These antibodies are called “unexpected”

because only minor part of general population
have positive antibody screen.
• Uses of detection and identification of red blood cell antibodies –

 For the selection of appropriate blood for transfusion

 Hemolytic disease of the new born

 Immune hemolytic anemia.

• It involves testing patient’s serum against screening cells

• Screening cells are available commercially or prepared in blood bank

lab- using group O cell suspensions (O +ve or O -ve) obtained from
individual donors.

• Group O cells are used so that naturally occurring anti-A or anti-B will
not interfere with detection of unexpected antibodies.

• The cells are selected so that the following antigens are present on at
least one of the cell sample;

D, C, E, c, e, M N, S, s, P, Lea, Leb, K, k, Fya, Fyb, and Jkb.

PREPARATION OF CELL PANEL FOR
ANTIBODY SCREENING AND
IDENTIFICATION
• Take samples of O group blood from regular voluntary
donors in CPD/CPDA.

• Prepare 2-4% cell suspension of all samples in saline.

• Set up rows for 2 test tube racks A & B.

• Add 1 volume of following antisera in pre-labelled test tubes

in rack A i.e. anti N, anti P1, anti Lea and Leo .
• Similarly add following anti-sera in pre-labelled tubes in rack B
i.e. anti- C, D, E, c, e, S, s, Fya ,Fyb, K, k , Lua , Lub

• Add 1 volume of cell suspension in all tubes in both A & B

racks.

• Incubate rack A at room temperature and rack B at 37˚C for 1

hour.

• Look for agglutination in all tubes of rack A and record the

results.
• Wash all the tubes of rack B three times with normal saline and
decant the last wash completely.

• Add 2 drops of AHG reagent. Centrifuge at 1000 rpm for 1

minute and look for agglutination. Record the result.

INTERPRETATION

• If there is agglutination with anti - sera, It indicates presence of

corresponding antigen on red cells.

• These cells can then be used for antibody screening.

 METHODS FOR ANTIBODY IDENTIFICATION

Before proceeding to antibody identification it is useful to check

the following:

• Medical diagnosis

• History of pregnancy, transfusion, reason for transfusion and

drug therapy including Rh immunoglobulin.
Then the following techniques are carried out:

• Saline room temperature technique

• Enzyme test

• Indirect antiglobulin test

• Albumin technique at 37˚ C

 SALINE ROOM TEMPERATURE TECHNIQUE
This test is used to identify (Ig M) cold reacting antibodies.

e.g. anti-M, anti-N, anti Lea anti І, anti P etc.

Test Procedure

1.Put 2 drops of patient serum in each tube.

2.Add 1 drop of panel cells, pt’s cell suspension(auto control) and

O cord cells to appropriate tubes.

3.Mix and incubate at 20-25˚ C for 1 hour.

4. Shake the tubes gently and look for
agglutination/hemolysis.

5. Record the results.

• Lowering the temp. to 4˚C during the test enhances

reaction of cold antibodies.
 ENZYME TEST

• This test is used to identify warm reacting antibodies (Ig G)

type.

• It enhances reaction of: Rh, Lewis and Kidd antibodies

• But it may weaken or inactivate certain antigens i.e. M, N, S,

Fya & Fyb.
 GRADING OF THE REACTIONS

Agglutination reactions are routinely graded as:-

• Negative (no agglutination).

• Weakly positive, and 1+ through 4+.

Degree of the positive reaction : indicates the amount of Ab

present not its significance.
COMPONENT SEPARATION
&
PREPARATION
 TYPES OF COMPONENTS

• Cellular components

• Plasma components

• Plasma derivatives
 CELLULAR COMPONENTS

• Red cell concentrate

• Leucocyte reduced red cells

• Platelet concentrate

• Leucocyte reduced platelet concentrate

• Platelet apheresis
 PLASMA COMPONENTS

• Fresh frozen plasma

• Cryoprecipitate

• Cryo-poor plasma
 PLASMA DERIVATIVES

• Albumin 5% and 25%

• Factor VIII concentrate

• Fibrinogen

• Immunoglobulins
APHERESIS
 APHERESIS(HEMAPHERESIS)

• Derived from greek word Aphairesis

• Means taking away or separation.

• Hemapheresis is removal of whole blood from donor and separation into

components

1.In it desired component is retained

2.Unwanted component and remaining constituents is returned to donor

 INDICATIONS

• Collect component for transfusion purpose

1. Platelets

2. Leucocytes

3. Plasma

4. Peripheral blood stem cells

• To remove pathological components from blood that is theraupeutic apheresis

 METHODOLOGY OF APHERESIS

• Manual method

• Apheresis machines

a) Intemittent flow centrifugation.

b) Continuous flow centrifugation.

 ADVANTAGES OF MANUAL METHOD

• Process is simple and inexpensive

• No sophisticated equipment is required

• Amount of component prepared per procedure is less than

automated devices.
 DONOR SELECTION FOR
APHERESIS PROCEDURES

• Donors should be normal healthy individuals and should meet the

standards set for blood donation.

• Interval between apheresis procedures should be at least 48 hours

and the amount of red cell loss should not exceed 25ml/week.

• Platelet apheresis should have adequate platelet count

>1,50,000/ul.
• Plateletapheresis donor must not have taken aspirin or any
drugs which affect platelet function during the last 48 hours.

• Frequent cytapheresis donors should be tested for plasma

proteins and anaemia.
• Serum protein concentration should be above 6.0gm/dl

• In case of granulocytopheresis the donors are given steroids or

sedimenting agents example hydroxyethyl starch to increase
granulocyte yield but care is to be taken not to introduce such
drugs in donors having history of hypertension, diabetes and
gastrointestinal ulcer
• Donors should be screened prior to apheresis procedure for all the
legally mandatory infectious disease markers for whole blood
donation

• Tests are HIV ,HBsAg and anti HCV should perferably be performed
by ELISA technique except in emergency situations when screening
of the sample can be done by rapid test device.
 PLATELETPHERESIS

• Removal of platelets from a donor with the return of donor red cells, white
cells and plasma.

• Yield of platelet depends upon donor initial platelet count and amount of
blood processed

• It takes 90 to 120 minutes for the process to occur

• Platelets can be stored for 5 days on platelet agitator with incubator at 22°C.
• If platelet is prepared in an open system it must be transfused
within 24 hours.

• Day of collection is counted as zero.

 LEUCOPHERESIS

• Removal of white cells with the return of red cells , plasma and
platelets to the donor.

• Base on the estimated blood volume of 7% of the body weight ,

it is calculated that a dose of 1.4x1o8 granulocyte/kg is required
to produce a granulocyte increase of 1000/ul.
• Granulocyte concenterate must contain a minimum of 1.0 x 1010
granulocytes.

• Product should be transfused as soon after collection.

• Maximum shelf life is 24 hours.

 ADVERSE EFFECTS OF APHERESIS IN
DONORS

• Main effect is attributed to citrate toxicity

• Normally citrate gets metabolized in the body but if amount exceeds donor may feel
numbness or tingling sensation around the mouth.

• There may be febrile allergic reactions, headache, mild hypertension and edema of
extremities.

• Low molecular weight HES may be safer alternative

• Hypocalcemia
 PLASMAPHERESIS / PLASMA
EXCHANGE

• It is procedure in which whole blood is withdrawn from donor.

• Prevented from coagulation upon withdrawal with the return of separated cellular
components to donor.

• If relatively small amount of plasma example 500 ml are removed and replaced by
normal saline the term plasmapheresis is used.

• If more plasma is removed then it becomes necessary to infuse plasma or plasma protein
fraction to replace lost plasma proteins this is called plasma exchange.
 INDICATIONS OF PLASMAPHERESIS

1. Removal of antibodies

2. Removal of immune complexes

3. Hyperviscosity syndromes

4. Removal of toxins

5. Replacement of deficient plasma components or immune

complexes
 COMPLICATIONS OF PLASMA EXCHANGE

• Reactions to replacement fluids

• Vasovagal reaction

• Pyrogenic reactions

• Hypocalcemia

• Thrombocytopenia

• Anaemia

• Hypogammaglobulinaemia
 PHOTOPHERESIS

• Photopheresis or extracorporeal photopheresis (ECP) is a form of apheresis in which

cell is treated with photoactivable drugs which are then activated with ultarviolet
light.

• It is currently standard therapy approved by the U.S. for cutaneous T – cell

Lymphoma.

• Also used in treatment of other T cell mediated disorders, including chronic graft
versus host disease and solid organ transplant rejection.
• Photopheresis is used for the treatment of autoimmune disease,
such as systemic sclerosis and rheumatoid arthritis.

• Process is performed through one intravenous access port and 3

basic stages

1.Leukapheresis

2.Photoactivation

3.Reinfusion
 MECHANISM OF ACTION

• The combination of 8-methoxy psoralen and UVA radiation causes

apoptosis of the treated T cells and may cause preferential
apoptosis of activated or abnormal T cells thus targeting the
pathogenic cells of CTCL or GVHD.
 PROCEDURE

1. Peripheral intravenous line or central venous access is

established in the patient.

2. Blood (225ml) is passed through three cycles of leukapheresis or

125ml of blood is passed through 6 cycles depending upon
patient hematocrit value and body size.
3. Collected wbcs are mixed with heparin , saline and 8-
methoxypsoralen

4. The mixture is passed as a 1mm film through asterile cassete

surrounded by UVA bulbs for 180 minutes

5. Treated WBC are returned to patient

 ADVERSE REACTIONS

• Hypotension

• Syncope

• Low grade fever

 DETECTION OF
TRANSFUSION
TRANSMITTED INFECTION
 ELISA
ENZYME-LINKED IMMUNOSORBENT ASSAY
• ELISA is the first test developed for the detection of HIV antibodies
in 1985.
• Currently most widely used test for serodiagnosis of HIV infection.
• ELISA detects substances with antigenic properties (mainly
proteins).
• Based on enzymatic color-reaction.
 BASIC PRINCIPLE OF ELISA

• Enzyme is used to detect the binding of Antibody – Antigen

• Enzyme converts colorless substrate into colored product,

indicating the presence of Antibody - Antigen complex

• ELISA can be used to detect either presence of Antigens or

Antibodies.
 TYPES OF ELISA

• Indirect Elisa

• Direct Elisa Non - Competitive Elisa

• Sandwich Elisa

• Competitive Elisa
 NUCLEIC ACID TESTING (NAT)

• Method of testing blood that is more sensitive than conventional

tests that require presence of antibodies or antigen to trigger a
positive test result.
 BENEFITS OF NAT

1. Detects low level of viral RNA or DNA

2. Highly sensitive and specific for viral nucleic acids

3. Provides additional layer of safety to the blood supply

4. Detects infection earlier than other screening methods,

narrowing the window period

5. Have the ability to detect viral mutants and occult infections.

 TECHNIQUES OF NAT

• Techniques for detection of viral DNA or RNA in donor/patient’s

blood sample. These include:

1. Polymerase Chain reaction (PCR)

2.Branched DNA (bDNA )

3.Transcription mediated amplification (TMA)

4.Nucleic Acid Sequence Based Amplification (NASBA)

• Power of NAT is its ability to detect the presence of
infection by directly testing for viral genomic nucleic acids
rather than by indirectly testing for the presence of
antibodies.
 NAT TECHNOLOGIES FOR BLOOD
SCREENING

• FDA approved NAT assays

for blood screening are PCR
and TMA.
 COMPARISON
ISSUE TMA PCR
Technology •Transcription mediated amplification •Polymerase chain reaction (PCR)
(TMA)
•Amplification products: RNA •Amplification products: cDNA
Amplicons. Amplicons.

•Isothermal reaction, wherein •Requires a thermal cycler

everything occurs at the same instrument to rapidly change the
temperature in a water bath temperature for amplification
reaction.

•TMA produces 100-1000 copies per •PCR doubles the copy number per
cycle. This results in a 10 billion fold cycle
increase of copies within about 15-30
minutes.
Sample •No need of ultra centrifugation , special •Ultracentrifugation is required,
preparation sample preparation and extraction sample preparation needs many
steps.
•Technician may miss the pellet
•Requires >13 manual steps and 2
manual sample transfers

•Convenient procedure minimizes the •Chances of mistake increases

chances of mistake by technicians due to the complexity of the
procedure
Additional sample •No sample preparation •Manual step
preparation issues •No risk of false negatives as target nucleic •Risk of false negative results due
acids are seperated using a magnetic field to “sucking”pellets.
•Low risk of contamination (RNA amplicon) •High risk of carryover and /or
•Traceability throughout the process as the contamination (double- stranded
whole process is performed on the same amplicon)
barcoded tubes with no transfers •Lack of traceability due to the
manual transfer of sample
material and eluate.
Sample monitoring •Internal control is being added in •This facility is not available for
facility all the tubes to monitor the assay each type of tests.
performance of all the samples
separately

Contamination issue •All the steps are performed in a •Transferring of samples from one
single tube tube to another during extraction
and detection
•This minimizes the chances of •It may lead to contamination
contamination
IDT Vs pooling •Individual donor testing (IDT). •Pooling results in a dilution of
•IDT has higher sensitivity viremic samples proportionate to
the pool size which reduces the
sensitivity.

•Faster release of blood and blood •Resolution of reactive pools will

products. add cost, time and manpower
ADVANTAGE OF TMA FOR NAT TESTING

1. Amplification product-RNA Amplicon. RNA is more labile in the

laboratory environment than DNA , this helps reduce the
possibility of contamination

2. Isothermal reaction: amplification occurs at the same

temperature in a water bath

3. TMA produces 100-1000 copies per cycle (this results in 10

billion fold increase of copies within 15-30 minutes
4. No need of ultra centrifugation, special sample preparation and
extraction.

5. Internal control monitors for each sample from start to end

separately

6. All the steps are performed in a single tube minimizing

contamination chances.

7. Presence of heparin does not inhibit the reaction

 POTENTIAL EFFECT OF NAT

Disease Window period

(days ) (days)
HIV 21 days 6 days
HBV 36 20
HCV 65 3
Benefits of ID-NAT over Minipool NAT Testing
• IDT NAT : samples tested individually without any dilution
• Highest Sensitivity: maintains greatest level of sensitivity without
any dilution of viral genetic material prior to testing
• Higher throughput: Streamlines operational process without any
pooling steps.
• Faster release: Ensures zero delay in releasing negative sample
results without resolution of reactive pool.
• Less chances of contamination.
 Minipool testing is even less effective for window period HBV
detection as the viral load is often low
 Worldwide there is no known or published case of transmission
transmitted infection (TTI) after screening by ID-NAT while several
case have been reported in minipools.
 RECENT TECHNOLOGY
&
AUTOMATION IN BLOOD BANKING
QUANTIFICATION OF BLOOD GROUP A AND
B ANTIBODIES BY FLOW CYTOMETRY

• In clinical management of patients receiving blood group ABO-

incompatible organ allografts, it is of importance to determine the
levels of blood group A and B antibodies before and after
transplant.
• Currently used methods, which are mostly based on
hemagglutination, are improper and are associated with large
intercenter variations.
• A flow cytometry-based assay for the semiquantification of blood
group A and B antibodies
 DETECTION OF ABO BLOOD GROUP
POLYMORPHISM BY DENATURING
GRADIENT GEL ELECTROPHORESIS

• Use of polymerase chain reaction (PCR) format together with

denaturing gradient gel electrophoresis (DGGE) allows rapid
identification of the 6 major genotypes (AA, AO, BB, BO, AB and
OO) of ABO blood group polymorphism in a single amplification.

• Also distinguishes undescribed polymorphisms associated with

the A and B alleles.
• PCR-based methods have been described to allow the detection
of gene polymorphisms responsible for many blood group
antigens.

• These methods are routinely used to test samples of fetal origin

and to resolve serologic discrepancies.

• Typing of blood donors for minor antigens to facilitate the

procurement of compatible blood for alloimmunized patients.
• Allow multiplex amplification of specific fragments of blood group
genes and the fragments of the Rh (d, c, c, e, e), kell (k, k), duffy
(fya, fyb), and kidd (jka, jkb) genes could be amplified.
 DNA-ARRAY TECHNOLOGY

Hemagglutination being used in immunohematology reference

laboratory has limited capability to resolve complex problems
Limitations-
•Hemagglutination is subjective
•Does not reliably predict a fetus at risk of hemolytic disease of the
fetus and newborn (HDFN),
•Difficult to phenotype RBCs from a recently transfused patient or
when RBCs are coated with IgG
•Has poor ability to predict zygosity in Rh-positive individuals
• DNA array technology has the added advantage of computerized
interpretation and documentation of results, and direct
downloading to a database.

• Molecular testing using DNA arrays, makes it feasible to

contemplate mass screening donors to increase inventories of
antigen-negative RBC components, and precisely matching the
antigen-negative status of a transfusion recipient to that of a
donor.
• DNA analysis with BeadChip format, combined with computerized
data entry and analysis, permits the prediction of minor blood
group antigens.
AUTOMATION IN BLOOD BANKING
 DUET BLOOD GROUPING READER

A highly sensitive image analysis

reader for the interpretation of
blood grouping plates, gel cards
and other agglutination based
assays.
• Suitable for ABO/Rh grouping, antibody screening etc

• Reads all varieties of plates and gel type cards

• Fully programmable, high-resolution image analysis

• Detects and interprets "weak" positives reliably

• Batch mode option for maximum throughput

• Automatic bar-code reading of plates and cards.

COMPACT AUTOMATED BLOOD GROUPING SYSTEM

• The BeeGroup 300 is a compact and

flexible instrument.

• Cost-effective automation with

maximum possible security.

• The BeeGroup 300 automates all

pipetting, reagent addition,
incubation and washing steps.
The BeeGroup 300 can be used to perform all of the following tests:

• ABO Grouping

• Rh Typing

• Antibody screening

• Cross matching
• A range of different card and plate racks is
available to match reagent supplier's
consumables.

• Furthermore, the deck of the BeeGroup 300

has a unique racking system which allows
different layouts to be readily interchanged.
4 microplates only, 2 microplates and 12
cards, 3 microplates and 12 cards or 24
cards only.
CLINICAL UTILITY OF SCREENING DONORS FOR ALL
CLINICALLY SIGNIFICANT POLYMORPHISMS

• To provide fully matched blood

 Transfusion-dependant patients
 Women of child bearing age
• To provide compatible blood to patients with antibodies
• To perform full electronic matching
 Reduce alloimmunization
 Reduce DHTRs
 Reduce serological testing 203
THANK YOU