Blood Grouping and Rh Typing: Introduction, Principle, Procedure, Result- Interpretation, and Keynotes


Blood Grouping and Rh Typing are used to determine the blood group and Rh factor of a sample of blood. ABO blood grouping and Rh typing are the most common blood group systems. There are nearly 300 blood group systems so far discovered. Other common remaining systems are MNS, Lutheran, Kell, Duffy, etc. Karl Landsteiner discovered ABO (A, B and O) in 1900. Two years later, Decastello and Sturli added AB and finally ABO system gets completed. In 1940, Karl Landsteiner and Winner discovered Rh factor in red blood cells (RBCs) of the Rhesus monkey (Macca rhesus).

For his great contribution to blood banking and transfusion medicine, he was awarded the Nobel Prize in Physiology or Medicine in 1930. He is also known as the Father of Blood Grouping and Immunochemistry. Methods of ABO Blood Grouping and Rh Typing are of two types i.e. slide and tube. But here we are concerned with the slide method of ABO blood grouping and Rh typing as it is rapid, simple, and easy to proceed.


ABO blood grouping and Rh typing are based on the principle of agglutination. The normal red blood cells (RBCs) possessing antigens will clump in the presence of corresponding antibodies.

Test Requirements

  • Antisera A,B and Rhesus (Rh)
  • Clean and grease-free glass slides
  • Clean sticks for mixing (optional since slide can be used for this purpose)
  • EDTA blood or clotted blood/ skin puncture using lancet at spot test
  • Micropipette and tips/dropper-optional in case of skin puncture
  • Physiological saline (optional)


  1. Label the two different clean slides with the unique ID of the patient.
  2. Pipette 50/50 µL (one-one drop) of the blood and deliver it to the first slide in two corners and a drop in the remaining slide at the center.
  3. Put 1 drop of A, B, and Rh antisera onto each slide respectively, and mix it thoroughly with the help of the clean sticks or using a clean glass slide.
  4. Tilt the glass slide back and forth for up to 2 minutes.
  5. Look for the agglutination in each slide. 
  6. In the case of Rh, if agglutination is seen it is reported as Rh positive and if no agglutination is seen, it is reported as Rh negative as shown table of result-interpretation.
Blood Grouping and Rh Typing: Introduction, Principle, Procedure, Result Interpretation and Keynotes
Fig. Blood Grouping and Rh Typing Antisera and Result Interpretation

Result Interpretation

ABO Typing
Rh Typing
+ Rh Positive
Table of ABO and Rh typing Result-Interpretation

Applications of ABO Blood Grouping and Rh Typing

It is applicable for the following streams-

  • Prior to blood transfusion and cross-match.
  • In the case of hemolytic disease of the newborn (HDN)
  • Determination of paternity in medico-legal cases
  • Relationship of blood groups, and susceptibility to a variety of diseases e.g. blood group A is more prone to gastric carcinoma whereas blood group O is more prone to systemic lupus erythematosus (SLE).


  1. In the case of doubtful agglutination reactions, a further microscopic examination should be done to confirm the result.
  2. Each negative Rh result should be confirmed by the Du Test.
  3. For clarification of the agglutination, add a drop of normal saline/physiological saline. Tilt the glass slide back and forth and observe for agglutination/clumping.
  4. To omit prozone and post zone phenomenon, use 10% suspension of RBCs in place of using whole blood.

Further Reading


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