BLOOD GROUPING

BLOOD GROUPING

1. Who discovered the blood group system for the first time?

Ans: Karl Landsteiner

    Karl Landsteiner described ABO blood grouping in 1900 while Rh system was discovered later in 1940 by Landesteiner and wiener

2. What is Landsteiner Law?

• If an agglutinogen is  present on red cell membrane the corresponding agglutinin must be absent in the plasma

• If agglutinogen is absent on red cell membrane the corresponding agglutinin must be present in the plasma.

3. Name a few blood group systems

Ans: ABO system, Rhesus system, MNS system, Lewis system, Kelly, Duffy, Lutheran, Kidd system, Duffy system, Li and P etc.

4. What type of antibodies are found in blood group “A”?

Ans:   Blood group ‘A’ has anti -B antibodies in the serum

5. What type of antibodies are found in Blood group ‘B’

Ans:  Blood group ‘B’ has anti –A antibodies in the serum 

6. What type of antibodies are found in Blood group ‘O’

Ans:  Blood group ‘O’ has both Anti –A and Anti –B Anti –bodies in the serum

7. What type of antibodies are found in Blood group ‘AB’

Ans:  No antibodies

8. Which type of blood group individuals are known as universal donors

Ans :  Blood group ‘O’ individuals are known as universal donors

9. Who is a universal recipient?

Ans:  Blood group AB individuals

10. What are the methods used for blood grouping

• slide or tile method

• Tube method

• Microplate method

• Automated method

11. What is the procedure of Tile method for blood grouping?

        Procedure:

• Place 1 drop of anti A (blue) and 1 drop of anti B (yellow) reagent, and D reagent separately on a labelled slide or tile.

• Add 1 drop of blood to each drop of the type typing antiserum.

• The cells and reagent are mixed using a clean stick. Spread each mixture evenly on the slide or Tile over area of 10-15mm diameter.

• Tilt the slide (or) mix with stick and leave the test for 5 min at room temperature (22-24) and look for agglutination with naked eye .It is minimal –confirmed by microscope.

12. What is procedure of tube method for ABO grouping?

Cell grouping: In this method cells and serum of unknown blood samples to be tested.

• Prepare a 2-5% cell suspension in saline from unknown blood sample.

• Take 3 test tubes1, 2, 3 and put a drop of anti-A (blue), anti-B (yellow) and anti-AB serum (pink) to them.

• Add one drop of red cell suspension in cach test tube.

• Centrifuge at 1500rpm for one minute.

• Look for agglutination either with naked eye or under the microscope.

Serum grouping:

• Allow unknown blood sample stand for some time and separate the serum.

• Add2 drops of unknown serum in test tube4,5,6.

• Add 1 drop of 2-5% cell suspension of known blood of A,B and O group into these test tubes.

• Centrifuge at 1500rpm for one minute.

• Look for agglutination either with naked eye or under the microscope.

Results of ABO grouping

Blood group	Agglutination +with anti A	Agglutination with anti B
A	+	–
B	–	+
AB	+	+
O	–	–

 

13. What is the procedure of tube method for RH grouping?

• Prepare a 2-5% cell suspension in saline from the blood to be tested.

• Take a test tube and put a drop of anti-D serum

• Add one drop of red cell suspension in the tube

• Centrifuge at 1500rpm for 1 minute

• Look for agglutination either with naked eye or under the microscope.

14. For blood grouping which method is better tube or slide method

Ans: The tube technique is better and accurate because  

1. Weaker antigens like A2 also detected

2. Both red cell and serum grouping done.

 

15. What are the two ways for ABO blood grouping?

Ans: Cell grouping (forward or direct grouping) and serum grouping (indirect or reverse grouping)

16. What is cell grouping or forward or direct grouping

Ans:  Cell grouping or forward grouping includes the testing of antigens present on the RBC s of the patient with the help of different commercially available antisera.

17. What is serum grouping or reverse or indirect grouping

Ans: Serum grouping or reverse grouping is the testing of antibodies present in the serum of the patients with the help of known cells of different blood groups

18. What is cross matching?

Ans: It is direct matching of blood components (serum&cell) of recipient and donor red blood cells

19. What is cross matching procedure?

• In a small test tube, place a drop of recipient’s serum

• Add washed donor red cells suspended in 5% saline

• Mix the two and incubate at 37ºc for 30 minutes

• Centrifuge it at 3000rpm for one minute.

• After dislodging the cell palette gently from centrifuge tube, examine it for presence or absence of agglutination and haemolysis, first grossly and then under the low power of the microscope.

20. What is the purpose of cross matching or compatibility test?

Ans:  The test is to prevent transfusion reactions by detecting  Antibodies in recipient serum, to detect ABO incompatibility between donor and recipient

21. Uses of blood grouping

• For blood transfusion

• In paternity disputes

• Hemolytic disease of new born

• Medicolegal use

• Immunology, genetics and anthropology

• Susceptibility to various diseases Ex: O group- peptic ulcer

• Part of Health check-up and job, driving licensing

 

22.  What are the adverse effects of transfusion?

Immediate reactions:

• Immediate febrile reaction

• Acute hemolytic transfusion reaction(fever, chills, tachycardia)

• Allergic reactions(urticaria, itching, pruritus)

• Anaphylactic reactions(Rash, itching)

• Transfusion associated circulatory overload (TRALI)

• Bacterial contamination of blood

• Biochemical upsets following transfusion

• Transfusion associated lung injury(Fever, chills, dry cough)

Delayed reactions

• Delayed hemolytic transfusion reaction (fever, anemia)

• Transfusions of infections

• Iron over load

Graft vs host diseases

• Post transfusions purpura

23. What are all investigations that are performed on a donor before drawing the blood

Ans:  Hb estimation, Hepatitis-B, Hepatitis C, Syphilis, HIV, malarial parasite, ABO grouping, Rh-D typing and Antibody screening.

24. What do you mean by Rh Incompatability?

• Rh incompatibility occurs when Rh-ve mother has Rh + ve foetus.

• During first pregnancy foetal blood crosses the placenta in to mothers blood to produce Ig G antibodies.

• In subsequent pregnancies antibodies crosses later to foetus and cause immune destruction in fetal RBC

25.  What is Bombay Phenotype?

• The Bombay blood group was first discovered in 1952 by Bhende and others.

• The pheno type is characterized by the absence of A,B,H antigens on the red cells. The serum of the these persons contain anti A, anti B, and Anti H antibodies

26. What is coombs (Anti globulin) test

• It is used to demonstrate presence of Ig G antibodies against red blood cells in the patient

• IgG antibodies do not cause agglutination of RBCS, coombs serum is added which combines with IgG on RBCs surface resulting agglutination.

27. What is the procedure of direct coombs test?

Ans: Take patient’s RBCs from the clot/EDTA blood/cirated blood in a test tube and add normal saline and mix. Centrifuge at 2500 rpm for 5 minutes. Decant the saline.

• Wash RBCs twice more in the same manner

• After three washings, prepare a 5% saline suspension of RBCs

• Take 2 drops of saline red cell suspension and add 2 drops of anti-globulin serum and mix well. Incubate at 37degrees for 5 minutes. Centrifuge at 1000rpm for 1 minute.

• Dislodge the cell button, gently resuspended the cells and observe for aggluation both microscopically and macroscopically. With each test, one positive control and one negative control is set up.

• Positive control. Take 1ml Rh+ve EDTA blood and wash with N-Saline thrice. To 2 drops of 5% saline suspension and 2 drops of anti-Rh (anti-D) IgG antibody. Incubate at 37 degrees for 10 minutes for antibody to get coated on the surface of RBCs. Wash with saline thrice to remove the excess of Ab. Now these RBCs and 2 drops of coated RBCs and 2 drops of antiglobulin serum. Incubate at 37ºc degree aggluatination should be positive

• Negative control. Normal individuals RBCs- non sensitized RBCs serve as a negative control.

RESULT: Aggulation in the test sample indicates the presence of antibody on the surface of RBCs- Positive direct coombs test.

28. What is the procedure of indirect coombs test.?

Ans: This test is used to demonstrate antibodies that are present in the serum.

• Take 2 drops of patient’s serum in a test tube.

• Add 3 drops of 5% “O” cell suspension. These cells are taken from any “O” group person who is Rh+ve.

• Incubate the mixture at 37 ºc degree for 10 minutes.

• Wash with in-saline thrice to remove the excess of antibodies.

• Remove the suspernant.

• Now “O” RBCs are coated with (IgG) Anti-Rh antibodies of the patient (If antibodies are present).

• Add 2 drops of coomb’s serum (anti-globulin)

• Incubate at 37 degree for 10 minutes.

• Centrifuge at 2000 rpm for 1 minute

• Resuspended the red cells by gentle agitation and examine macroscopically and microscopically for agglutination.

Result: In the positive indirect coombs test, agglutination of RBCs indicates the presence of antibodies in the test serum.

29. What are direct combs test applications?

• Hemolytic transfusion reactions

• Auto immune hemolytic anemia

• Hemolytic disease of new born

• Drug induced hemolysis

30. What are indirect coombs test applications?

    Indirect:

•  Compatibility testing
•   Detection of unexpected antibody(IgG)s to Rh factor  in serum of pregnant patients
• Detection of red cell antigens not detected by other technique.

31 .What are blood components

Ans:  A Single unit of whole blood can be separated in to one unit of packed red cells, one unit of platelets and one unit of fresh frozen plasma

32. What is one unit of blood?

 ANS: 450ml of blood in 63ml of anticoagulant CPD (citrate phosphate-Dextrose) or CPDA (citrate phosphate-Dextrose Adenine).

33. What is the difference between complete and incomplete antibodies?

ANS: Naturally occurring anti A, anti B antibodies are IGM types- complete antibodies and cause agglutination in saline suspension.

• Incomplete antibodies IgG type and result because of antigenic stimulation and don’t cause agglutination in saline suspension.

34. What test is employed to detect low level antibodies against red cells?

ANS: Coomb tests.

35. Which are the immune haemolytic anemias in which coombs test is indicated?

ANS: Auto immune haemolytic anaemia SLE, cold agglutinin disease, paroxysonal small cold haemoglobinuria, infectious mononucleosis.

36. How are blood and its components stored?

ANS: Blood is stored in a refrigerator at 2-4 c upto 4-6 weeks, and platelet can be stored up to 5 days.

37. Which blood groups has subtypes?

Ans:  A has A1- the stronger antigen and A2 has- the weaker Ag. A2 Ag may sometimes be missed, if outdated antisera are used and the patient may be labelled as O. So, to prevent it, add anti- Ab serum to patient’s red cells. This would aggluniate all Ags except O. Therefore A2 antigen is detected.

38. Which are secretors?

Ans: A, B, and H antigens (A, B and H substances) can be found in the saliva of majority of people and are called “Secretors” while the rest are non-secretors.

39. What is the H gene and its role?

 Ans: Two main antigens A and B result in 4 blood groups- A , B, AB and O. These antigens are under control of A and B genes. Expression of A and B antigen appears to be dependent on product of gene H.

Interaction of H, A and B leads to:

•  H gene leads to secretion of basic precursor substance H.
• H substance is partly converted under influence of A and B genes into A/B substance, some of the H substance remains unconverted
• Since O group individuals do not have A/B genes, therefore no A/B substance is formed and these have large amount of H substance.

40. What are blood group antibodies?

Ans: Blood group antibodies are those antibodies which are produced against blood group antigens present on red cell surface. These are capable of reacting specifically with red cells of different blood group.

 

These AB antibodies are naturally occurring complete antibodies (IgM) and react strongly at temperatures lower than 37degree. These antibodies develop as a response to antigens that are similar to the human blood group and which are widely distributed in nature(horse serum, plant lectin, E coli bacteria, etc).Anti-A and Anti-B present in a new born infant at birth are passively acquired from mother in utero.

41. Is there any relationship between blood groups and susceptibility to disease?

Ans: There is a statistical evidence of such relationship but no casual relationship has been established yet.

• Blood group A—– Cancer of stomach, salivary gland, pancreas and pernicious anaemia.
• Blood group O- Duodenal ulcer
• Blood group Duffy- Susceptible to P vivax malaria.

42. How are the ABO antigens derived? Where are these found?

Ans: The basic substance of antigen is a cell membrane glycoprotein precursor which is converted to H substance under the influence of H gene (HH or Hh phenotype). The ABO genes are located at different loci. The A and B genes each produce a glycosyl tranferase that converts H substance into A OR B antigens. 0 is a silent gene that leaves H unchanged.

These blood group antigens are found on the red cell membrane, on the membranes of all vascular endothelial cells and also on some epithelial cells. There lies the importance of ABO matching with renal or other allografts.

43. What is Rh system? How is the antigen derived?

Ans: If red cells of rheus monkeys are injected in to other animals, an antiserum (antirhesus serum) is produced which will agglutinate the red cells of about 85% of human beings. These cells are designated as Rh positive cells. These rare individuals who lack Rh antigens (Rh null cells) suffer from spherocytosis and shortened life span of red cells. These are three very closely linked loci occupied by genes C, D, E and their alleles c, d, e. These genes are responsible for red cell antigens C, D, E. c&e.

44. What is massive blood transfusion?

Ans: massive blood transfusion is when more than the patient’s blood volume is transfused in 24 hours or a single transfusion of 2500ml done.

45. Name the different types of blood transfusion

• Indirect transfusion

• Autologous blood transfusion

• Marrow transfusion

• Direct from one person to another in energy in war field.

• Peritoneal transfusion.

46. What are the indications of blood transfusion?

• Haemorrhage

• Peripheral circulatory failure

• Severe infection

• Miscellaneous like severe anaemia, corrosive and radiation therapy.

47. What should be the age of human volunteers for blood donation?

Ans: 18 to 55 years with normal haemoglobin levels.

48. How much time is required for compensation of blood donors?

Ans: The time taken to complete compensation of loss of blood is approximately 3 months. So after 3 months a donor can repeat blood donation.

49. What is the locus genetics for blood group antigens

Ans: The genetic loci are situated mostly in autosomes.The locus for ABO group is on chromosome 9 and that for Rh group is on chromosome 1.

References

1. S.Sanyal. Pathology Practical book, 4th edition

2. Harsh Mohan. Practical pathology, Third edition with viva voice questions

3. Dr .Tejindar Singh,  Dr .K. Uma  Chaturvedi.Practical pathology for undergraduate and post graduate students-First edition

4. Ganga S. pilli. Viva in pathology, First edition

5. Internet sources

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