PART 2: Unit 6 Objectives

Why are ABO antibodies expected?

• they develop in response to environmental exposure

• the body forms antibodies against the ABO antigens it lacks

• they form naturally

Why do blood banks not routinely perform ABO reverse grouping on infants less than 4 months old?

• they do not develop antibodies till 4-6 months old

List the reagents routinely used to perform ABO forward and reverse groupings in the Lab.

• Forward

  • Anti-A

  • Anti-B

  • Anti-A,B

• Reverse

  • A1 cells

  • B cells

Why is the Anti-A,B reagent special in blood banking?

• it reacts with both A and B antigens on RBC

• used to:

  • confirm ABO typing

  • detect weak A or B antigens

  • can resolve discrepancies

  • gives very strong reaction with AB cells

Define ABO discrepancy.

• mismatch or unexpected reaction between the forward (cell) and reverse (serum) in ABO typing results

How do you identify an ABO discrepancy

1. perform forward type (RBC antigens)

2. perform reverse type (serum antibodies)

3. compare

   1. match: no discrepancy

   2. mismatch → ABO discrepancy

What are examples of ABO discrepancies?

• Forward

  • Anti-A: +

  • AntiB: -

• Reverse:

  • A cell +

  • B cell: +

What is an example of a Bombay pattern ABO discrepancy?

• Forward

  • Anti-A: -

  • Anti-B: -

• Reverse

  • A cell: +

  • B cell: +

State the normal expected ABO agglutination reaction strengths in a normal healthy adult in a forward typing test.

Blood Group Anti-A Anti-B

A 4+ 0

B 0. 4+

AB 4+ 4+

O 0 0

State the normal expected ABO agglutination reaction strengths in a normal healthy adult in a reverse typing test.

Blood Group Anti-A Anti-B

A 0 4

B 4+ 0

AB 0 0

O 4+ 4+

What is Whartons Jelly?

• gelatinous connective tissue within the umbilical cord that can interfere with ABO typing

How can Whartons Jelly Interfere with ABO typing?

• can coat RBC and make them sticky causing false positive agglutination

• can dilute plasma/serum and dilute antibodies causing weak reverse typing tests

• reduces newborn sensitivity to reverse typing

Define lectin.

• protein that binds to certain carbohydrate structures on cells or molecules without altering them enzymatically

• used in blood banking for:

  • detecting blood group antigens

  • identify weak antigens

What is the purpose of ISBT nomenclature?

• avoid confusion between different names for same allele or antigen

• provide unique numeric code for each antigen or allele

• standardize reporting in transfusion medicine

How does the ABO system work under ISBT representation?

System name: 001 (ABO system)

Antigens: each has a unique numeric code

Alleles: assigned specific allele numbers

How does ISBT nomenclature work?

• Lab reporting

  • does not say A or B instead uses ISBT code

• Genotyping and testing

  • genetic variants are assigned unique allele codes

  • helps avoid confusion

What are the different ABO blood compatibilities for transfusions.

Recipient Can Receive RBC from

O O only

A A or O

B B or O

AB A,B,AB, or O

How do you select which RBC units are safe for someone to have?

1. determine patient ABO type

2. check which antigens they do NOT have

   1. these are antigens that plasma antibodies will attack

3. select donor RBC that do NOT have these antigens

Given someones ABO type, what plasma transfusions can they receive?

Recipient ABO Type Antibodies in Plasma Can Receive Plasma

O anti-A and anti-B O,A,B,AB

A anti-B A, AB

B anti-A B, AB

AB none AB only

Why is AB plasma considered the universal donor plasma?

• it contains no anti-A or anti-B antibodies

Given a patients Blood Type, what compatible whole blood donor types would work?

Recipient Compatible Donors

O O only

A A only

B B only

AB AB only

What pre-analytical errors can affect ABO typing?

• mislabeling/wrong patient ID

• sample contamination/hemolysis

• clotted specimen

• delayed transport/improper storage

• collection from IV line with incompatible fluids

errors that lead to incorrect ABO type assignment or mixed-feild reactions

What analytical errors can impact ABO typing?

• incorrect reagent use

• expired or degraded reagents

• improper technique (shaking, timing temperature)

• failure to perform reverse typing

• reading errors

lead to incorrect results on the testing platform

What post analytical errors can impact ABO typing?

• transcription/ date entry errors

• failure to resolve ABO discrepancies

• mismatched labeling of blood units

• incorrect communication with clinical staff

Who discovered ABO blood groups?

• Karl Landsteiner

What year did Karl Landsteiner discover ABO blood groups?

• 1900 - 1903

Describe the formation of the H antigen

Allele product

• frucosyltransferase enzyme encoded by H (FUT1) gene

Action

• add L-frucuse to terminal sugar of type 2 oligosaccharide chain

Result

• formation of H antigen (foundation for A and B antigens)

Describe the formation of the A antigen.

Allele product

• N-acetylgalactosaminyltransferase encoded by A allele

Action

• transfer N-acetylgalactosamine into terminal fructose of H antigen

Result

• convert H antigen → A antigen on RBC surface

Describe the formation of the B antigen.

Allele Product

• Galactosyltransferase encoded by B allele

Action

• transfers D-galactose onto the terminal fucose of the H antigen

Result

• convert H antigen → B antigen on RBC surface

Where do the A,B, and H alleles perform the action to turn into antigens?

• golgi apparatus of erythroid precursor cells

What is Landsteiner’s Rule?

• a person naturally forms antibodies against ABO antigens that are absent on their own RBC

What are the ABO/H antibodies formed by each blood group?

Blood Group RBC antigens present naturally occuring antibodies

A A + H Anti-B

B B+H Anti-A

AB A+B+H none

O H only. Anti-A and Anti-B

Bombay none Anti-A Anti-B and Anti-H

What is the purpose of Forward Typing?

• identify antigens on the patients RBC

How is forward typing done in tube hemagglutination testing?

1. Prepare RBC suspension

   1. 2-5% suspension of patient RBC in saline

2. Add commercial antisera

   1. Anti-A reagent → detect A antigen

   2. Anti-B reagent → detect B antigen

3. Mix gently and centrifuge

   1. 60 seconds at 1000 rpm

4. Examine agglutination

   1. agglutination = antigen present

   2. no agglutination = antigen absent

How is reverse typing done in tube hemagglutination testing?

1. Prepare plasma or serum

What is the purpose of reverse typing?

1. Identify antibodies in the patients plasma/serum

   1. 2 drops of patient plasma

2. Add reagent RBC

   1. A1 red cells → detect anti-A antibodies

   2. B red cell → detect anti-B antibodies

3. Mix gently and centrifuge

   1. 60 seconds at 1000 rpm

4. Examine agglutination

   1. agglutination = antibody is present

   2. no agglutination = antibody is absent

What phase is forward typing performed at?

• perform at RT or cold sometimes confirm with immediate spin phase in routine ABO typing

• agglutination is IgM mediated

What phase is reverse typing performed at?

• cold or RT because ABO antibodies are mainly IgM

How do you use forward typing results to predict reverse typing?

1. identify antigens present on RBC

2. use Landsteiner’s Rule:

   1. Patient has antigen → no antibody against it

   2. Patient lacks antigen → antibody against it

How do you use reverse typing results to predict forward typing results?

1. identify antibodies present in patient plasma

2. predict which antigens are present on patients RBC

   1. patient has antibody → RBC lacks that antigen

   2. patient lacks antibody → RBC has that antigen

Give a Table showing the reverse typing results and predicted forward typing results.

Reverse Typing Predicted Forward Typing

Anti-A reacts RBC has B antigen → forward (Anti-B agglutinates)

Anti- B reacts RBC has B antigen → forward (Anti-A agglutinates)

Anti-A and B react RBC have no A or B → forward (no agglutination)

Neither react RBC have A and B → forward (anti-A/B agglutinate)

Give a Table showing the forward typing results and predicted reverse typing results.

Forward Typing Predicted Reverse Typing

Agglutinate: Anti-A Anti-B reacts (plasma has anti-B)

Agglutinate: Anti-B Anti-A reacts

Agglutinate: Anti-A and B neither reacts (AB)

No agglutination O Anti-A and Anti-B react

Forward Typing shows agglutination with Anti-A only, What does this mean?

• A antigen is present

• reverse typing plasma should contain Anti-B

Revere typing shows anti-A and anti-B in plasma, what does this mean?

• both anti-A and anti-B antibodies are present

• no A or B antigens are present

• forward test should not agglutinate with Anti-A or Anti-B

• blood type is O