The Blood Transfusion Laboratory

Question 1

What is present on the surface of red blood cells?

Answer 1

Antigens

Question 2

Where are antibodies found and what are they?

Answer 2

• Antibodies are found in the blood plasma
• They are immunoglobulins produced by the immune system following exposure to a foreign antigen

Question 3

What immunoglobulin antibodies are there present in the blood?

Answer 3

IgG (RhD) and IgM (Anti-A and Anti-B)

Question 4

When does a blood reaction occur?

Answer 4

When an antibody in the plasma reacts with an antigen on cells

Question 5

What is the effect of antigens in transfused blood?

Answer 5

• Foreign antigens in transfused blood will stimulate a patient to produce the antibodies only if the patient lacks the antigen (non-self)
  
  • The frequency of antibody production is very low, this frequency will increase as more transfusions are given

Question 6

What will stimulate antibody production? (3)

Answer 6

• Blood tranfusion
  
  • Blood carrying antigens foreign to patient
• Pregnancy
  
  • Foetal antigen entering maternal circulation during pregnancy
• Environmental factors
  
  • Naturally acquired e.g Anti-A and Anti-B

Question 7

What are the two types of antibody-antigen reactions?

Answer 7

• in vivo (in the body)
  
  • leads to destruction of the cell (e.g transfusion causing haemolysis) either
    
    • directly = cell breaks up in bloodstrem (intravascular)
    • Indirectly = liver and spleen remove antibody coated cells (extravascular)
• In vitro (laboratory)
  
  • Seen in agglutination tests

Question 8

What is agglutination?

Answer 8

The clumping together of red cells into visible agglutinates by antigen-antibody reactions to to the cross-linking of antibodies with antigens

Question 9

What can agglutination identify?

Answer 9

• The presence of a red cell antigen
  
  • I.e blood grouping
• The presence of an antibody in the plasma
  
  • Antibody screening/ identification

Question 10

What is the clinical significance of the ABO grouping system?

Answer 10

• If you transfuse red cells without knowing the patients blood group the chance of an interaction will be quite high
• ABO antibodies can active complement and cause INTRAVASCULAR HAEMOLYSIS

Question 11

What is the inheritance of blood groups?

Answer 11

• A and B genes are dominant
• O is recessive
  
  • 2 chromosomes are inherited (one from each parent) x 3 alleles = 6 possible combinations
    
    • AA, AB, BB, BO, OO, AO

Question 12

What does co-dominance mean?

Answer 12

Co-dominance means when both alleles are equally expressed and hence both contribute to the phenotype of the heterozygote.

This happens when AB genotype as they are both dominant

Question 13

If you have the OO genotype what red cell antigens and antibodies will you have?

Answer 13

• Phenotype will be blood group O
• You will have no red cell antigens
• You will have anti-A and anti-B antibodies

Question 14

If you have the AB genotype what red cell antigens and antibodies will you have?

Answer 14

• You will have the AB phenotype
• You will have A and B antigens
• You will have no red cell antibodies

Question 15

How do we find out the blood group of a particular patient?

Answer 15

Patients red cells and plasma are both tested

• This allows us to work out what antigens are on the surface of the red blood cells and what antibodies are present in the plasma

Question 16

How do we test a patients red cells to find out the presence of antigens?

Answer 16

• The gel matrix will be impregnated with anti-A, anti-B and anti-D antibodies
  
  • Agglutination will show that there is a particular antigen on the red cells
  • No agglutination shows the antigen is absent
    
    • Red cells will pass through the gel matrix, unless there is agglutination = big agglutinate will not pass through and stay at the top

Question 17

What would the blood type be of this person where we have added red blood cells to tubes with anti-A, anti-B and anti-D (forward) and plasma to the tubes with red blood cells (reverse)?

Answer 17

Blood group A as there is agglutination when the red cells were added to the anti-A test tube

If it were AB = agglutination in both, if it were O = no agglutination in the wells, no antigens present

Question 18

How do we test a patients plasma to find out the presence of antibodies?

Answer 18

• Gel matrix impregnated with A cells and B cells
  
  • Agglutination shows that a particular antibody is in the plasma or serum
  • No agglutination shows that the antibody is absent

Question 19

Explain why blood group O is considered the universal donor

Answer 19

• If you are blood group O you will have neither A nor B antigen
• Therefore, whatever antigens are in the recipients plasma there will be no interaction and agglutination
• Blood group O can donate blood to all blood groups

Question 20

Explain the Rh grouping system

Answer 20

• Second most important after ABO
• People with the D antigen are RhD positive (85% of UK)
• People who do not produce any D antigen are RhD negative
  
  • 4 main others are C,c,E and e

Question 21

What is the clinical significance of the Rh antigen in transfusion reactions?

Answer 21

• D antigen is very immunogenic and anti-D is easily stimulated = PREVENTION
• All Rh antibodies are capable of causing a severe transfusion reaction - ANTIBODY DETECTION

Question 22

What is the clinical significance of the Rh antibodies in pregnancy?

Answer 22

• Rh antibodies are usually IgG and can cause haemolytic disease of new-borns

Question 23

Explain haemolytic disease of newborn (HBN)?

Answer 23

1. Rh+ father so foetus in Rh- mother is Rh+
2. Rh antigens from foetus can enter mothers blood during delivery
3. In response to foetal Rh antigens, mother will produce anti-Rh antibodies
4. If a women becomes pregnant with another Rh+ foetus her anti-Rh antibodies will cross the placenta and damage foetal RBCs

Question 24

How can we avoid haemolytic disease of newborn?

Answer 24

• Carry out blood group and antibody screening at antenatal booking to identidy pregnancies at risk of HDN
  
  • RhD negative women may need anti-D prophylaxis
• Blood group and antibody screen at 28 weeks
• Atypical antibodies are quantified periodically to asses their effect in the foetus

Question 25

What treatment can we give a mother if she has identified as Rh-D and is carrying RhD+ foetus?

Answer 25

**Routine Antenatal Anti-D prophylaxis** Injection of anti-D to bind and remove any foetal RhD+ in the maternal circulation

Question 26

When is the anti-D injection administered?

Answer 26

* 1500 iu of anti-D is given routinely at 28 weeks and a smaller dose (usually 500 iu) after delivery if baby RhD+ * In some hospitals 2 smaller (500 iu) doses are given at 28 and 34 weeks instead of the 1 larger dose * Anti-D is also given after any event which may cause a feto-maternal haemorrhage (bleed between mum and fetus) such as: * Abdominal trauma * Intrauterine death * Spontaneous or therapeutic abortion

Question 27

How is antibody screening carried out for other clinically significant antibodies that can cause a haemolytic reaction?

Answer 27

* To detect for these antibodies * Patients serum is mixed with 3 selected screening cells containing relevant antigens, incubated for 15 minutes at 37 degrees + centrifuged * Clinically significant ABs reacting at body temp are detected and identified using a panel of phenotyped red cells

Question 28

Why is it important to screen and identify antibodies that cause haemolytic reactions?

Answer 28

It is important that we screen for these antibodies so that if detected, antigen negative blood can be provided to avoid stimulating an immune reaction

Question 29

What is the zeta potential?

Answer 29

Posotively charged ionic cloud that surrounds red blood cells

Question 30

What is the issue of this zeta potential (posotively charged ionic cloud surronding RBCs)?

Answer 30

* Red cells are not able to come in close proximity with the zeta potential * E.g pentameric IgM (anti-A and anti-B) antibody diameter allows agglutination but IgG antibody diameter too small to allow agglutination * For the IgG antibodies we need to get rid of the ionic cloud

Question 31

How we get IgG antibodies to interact with cells and remove the zeta potential?

Answer 31

LISS (Low ionic strength saline) * This will counteract the zeta potential

Question 32

What is the indirect anti-globulin test (IAT) used for?

Answer 32

* Use for * Screening for antibodies (IgG) * Identifying antibodies * Cross-matching donor blood with recipient plasma when there are known antibodies or a previous history of antibodies

Question 33

What is cross-matching?

Answer 33

Testing for agglutination of the donors RBC's by the recipients serum and vice versa

Question 34

What are the types of cross-matching we can do?

Answer 34

* **Immediate spin crossmatch (ISX)** * Must have had a negative antibody screen * Checking donor red cells against patient’s plasma * ABO check * Incubate for 2-5 minutes room temperature, spin and read * **Full Indirect Antiglobulin test (IAT) crossmatch** * Antibody screen positive or patient has known antibody history * Select antigen negative donor red cells and incubate with patient serum for 15 minutes at 37 degrees

Question 35

How do we test donor blood?

Answer 35

* Blood Establishment: * MHRA licensed manufacturer of blood and products * Donor Selection * Questionnaire: lifestyle, health, not previously transfused * Collection procedure arm cleansing / diversion pouch * **Comprehensive testing of all products** * **Viral** * HIV 1+2 * Hepatitis B * Hepatitis C * Syphilis * HTLV * **Platelets** * Bacteria * ABO, RhD, K, antibody screen

Question 36

What are the blood components for donation?

Answer 36

* Red Cells * Fresh Frozen Plasma (given for coagulopathy + associated bleeding) * Platelets (used to create clots to reduce bleeding) * **Cyroprecipitate** (contains factor VII, vWF and fibrinogen, used for peoples whos blood doesnt clot properly for example Haemophilia or vWdisease)