IMH LEC - LESSON 2: Relationship between Immunology & Immunohematology

Genetically encoded antigen system on the Erythrocytes, Leukocytes, Thrombocytes & Plasma

Blood group antigens

Enumerate the characteristics of Blood group antigens

1. Glycolipids (ABO)
2. Glycoproteins (Lewis Antigen)

Term for macromolecules capable of triggering an adaptive immune response by inducing the formation of antibodies

Immunogen

Term for small molecules that stimulate an immune response & is a partial antigen

Hapten

Enumerate the parts of an Immunogen

1. Hapten
2. Carrier

Enumerate the adaptive responses of Immunogens

1. Induce formation of antibodies
2. Sensitize T cells in an immunocompetent host

What complex determines the Human Leukocyte Antigen (HLA)?

Major Histocompatibility Complex (MHC)

Most Immunogenic Blood Group

A, B

2nd most Immunogenic Blood Group

Rh, D

If you have D antigen = ?

Rh positive (+)

If you DON'T have D antigen = ?

Rh negative = (-)

Enumerate the Immunogenicity of Blood Antigens

1. A, B
2. Rh, D
3. Kell (K)
4. Duffy (Fya)
5. Common Rh antigens

A foreign substance that reacts with antibody

Antigen

How long before a Transfusion reaction occurs with an incorrect blood type?

10-15 minutes

Antigenic property; Exact binding site of antigen to antibody

Epitope

Where can Epitopes be found?

Antigen

Exact binding site of antibody to antigen

Paratope

Where can Paratopes be found?

Antibody

Enumerate the Traits of an Immunogen

1. Macromolecular size
2. Chemical composition and molecular complexity
3. Foreigness
4. Ability to be processed and presented with MHC molecules (Antigen presentation)

Type of Epitope that consists of sequential amino acids on a single polypeptide chain

Linear Epitope

Type of Epitope that results from the folding of 1 or more polypeptide chains

Conformation Epitope

Term for an antigen that occurs in less than 1% of the population

Low incidence

Term when an antigen occurs in 99% of the population

High incidence

Products of antigenic stimulation and that demonstrate specific antibody activity

Antibodies/Immunoglobulins

AKA Gamma globulins

Antibodies/Immunoglobulins

What bond is present between chains in the antibody?

Disulfide bond

Blood Group Immunoglobulin that reacts with an antigen on the patient's own cells

Autoantibody

Blood Group Immunoglobulin that reacts with foreign antigen not present on the patient's own erythrocytes

Alloantibody

Enumerate the 2 types of Alloantibodies

1. Natural
2. Product of Immunization

What is the product of immunization to foreign erythrocyte by exposure through transfusion or pregnancy?

Immune antibodies

Enumerate Immunoglobulin classes from most predominant to least? (GAMDE)

1. IgG
2. IgA
3. IgM
4. IgD
5. IgE

Enumerate the 2 types of monomeric antibodies that can be produced from a polymeric antibody

* 2-mercapthoethanol (2-ME)
* Dithiothreitol (DTT)

Effect of Papain in a monomer

Cut monomer exactly at the hinge region and into 3 parts

Effect of Pepsin in a monomer

Cut monomer below the

Enumerate the factors that influence Antigen-Antibody reaction

1. Antigen-Antibody Ratio
2. pH
3. Temperature
4. Immunoglobulin type
5. Centrifugation

Effect to Ab-Ag ratio if it is a Homozygous concentration

Strong Ab-Ag reaction

Effect to Ab-Ag ratio if it is a Heterozygous concentration

Weak Ab-Ag reaction

Optimal pH level for Ag-Ab reaction

6.5-7.5

Temperature used to detect IgG

37C

Temperature used to detect IG

Room temperature

Reagents that enhance the detection of IgG antibodies by increasing their reactivity

Enhancement medium/ Potentiator

Substance that increases dielectric constant which reduces zeta potential of RBC

Protein media

Most commonly used protein media

* 22%/33% Bovine Serum
* Polyethylene Glycol (PEG)

Potentiator that removes water molecules in the environment

Polyethylene glycol (PEG)

Incubation time of PEG

15-30 minutes

Most commonly used Potentiator

Low Ionic Strength Solution (LISS)

Incubation time for LISS

5-15 minutes

Potentiator that targets the sialic component

Proteolytic enzymes

Test that detects antibodies and complement proteins that have attached to the RBC

Antihuman Globulin test

What animal is used to inject human serum into to produce antihuman serum?

Rabbits

Enumerate the types of Antiglobulin Tests

* Direct Antihuman Globulin (DAT)
* Indirect Antihuman Globulin (IAT)

Antiglobulin test that detects in vivo sensitization of RBC with complement components

Direct Antihuman Globulin (DAT)

Antiglobulin test that detects in vitro sensitization of RBC

Indirect Antihuman Globulin (IAT)

Enumerate the factors that affects AHG Test

1. Cell : Serum ratio (1:30)
2. Reaction medium
3. Temperature
4. Incubation time
5. Washing
6. Centrifugation (1,000 RCF for 20 mins)

Enumerate AHG Test Reagents

Polyspecific AHG, Monospecific AHG

Causes of False-Positive AHG Test results

* Over centrifugation
* Direct agglutination by strong cold agglutinins
* Over incubation with enzyme treated cells
* Improper use of PEG or polycation enhancement reagents
* Inadequate resuspension of cell button
* Rouleaux formation
* Dirty glassware
* Small fibrin clots may traps cells and mimic agglutination
* Cells with a positive DAT will yield false-positive results in any indirect antiglobulin test

Causes of False-Negative AHG Test results

* AHG reagent failure
* Failure to add AHG reagents
* Improper or inadequate washing
* Delayed washing
* Cell : Serum ratio too low
* Failure to add test serum or enhancement reagents
* Under centrifugation
* Resuspension of cell button too vigorous

Enumerate & differentiate the 3 positive reactions in Blood Bank

1. Agglutination
2. Hemolysis
3. Precipitation

Agglutination

Antibody-mediated clumping of particles that express antigen on the surface

Hemolysis

Rupture of Red cells with release of intercellular hemoglobin

Precipitation

Formation of insoluble complex from soluble antigen-antibody reaction

Appearance of RBC like a stack of coins

Rouleaux