ABO and Rhesus Blood Grouping principles and Result interpretation

Blood Grouping Kits MLTGEEKS

Blood grouping is a haematological haemagglutination test done to determine the blood group and Rh factor of an individual.It is the classification of blood based on the absence or presence of two inherited antigenic substances on the surface of red blood cells (RBCs) of the individual.

The ABO and Rh grouping are the major, clinically significant and the most important of all the blood group systems. Human ABO blood group system is divided into the four major groups depending on the type of antigen present on the surface of the red blood cells:

  • Blood Group A
  • Blood Group B
  • Blood Group AB
  • Blood Group O

ABO Blood Grouping System chart

Antigens on the surface of Red Blood Cells Antibodies in the Serum ABO Blood Group go site Genotype
AAnti B AAA or AO
BAnti ABBB or BO
A and BNo antibodiesABAB
No AntigenAnti A, Anti B, Anti AB OOO

These associated Anti A and Anti B antibodies usually belong to the largest immunoglobulin that is IgM class.

As for the  Rhesus system (Rh),It  is the second most important blood group system in humans.  Rhesus D antigen  is the most significant and immunogenic Rhesus antigen.Individuals carrying the Rhesus antigen are considered to have Rhesus positive blood group whereas those individuals lacking this antigen are considered to have Rhesus negative blood group.  

Principle of ABO and Rhesus Blood Group

The ABO and Rh blood grouping system is based on haemagglutination reaction. When red blood cells carrying either one or both antigens are exposed to corresponding antibodies leading to an interaction with each other to form visible agglutination or clumping.

Also read  WBC Count : Principle, Procedure and Calculation

ABO blood group antigens are O-linked glycoproteins in which their terminal sugar residues exposed at the cell surface of the red blood cells determine whether the antigen is A or B.

  • Individuals with blood A antigens on RBCs have anti-B antibodies in serum.
  • Those with blood group B have B antigens on RBCs and anti-A antibodies in their serum.
  • People with blood group AB have both A and B antigens on RBCs but lack anti-A nor anti-B antibodies in serum.
  • Blood group O individuals have neither A antigens nor B antigens, but possess both anti-A and anti-B antibodies in their serum.
  • As for the Rhesus antigens ,It is transmembrane proteins in which the loops exposed on the surface of red blood cells interact with the corresponding antibodies.

Blood grouping reagents

The blood Grouping reagents enables rapid identification of ABO blood group and Rh factor depending upon the antigen present on the surface of red blood cells.

Blood Grouping reagent
Blood Grouping reagent
Reagents go Color VolumesStorage temperature
Anti A Sera Blue5 ml 2-8 °C
Anti B Sera Yellow5 ml 2-8 °C
anti-ABClear5 ml 2-8 °C
Anti RhD Sera Clear5 ml 2-8 °C

Material Required for Blood Grouping

  • Antisera Reagents
  • 70% Alcohol or spirit
  • Cotton
  • Prinker or lancet
  • Mixing stick or applicator stick
  • Cavity slide or blood group slide

Precaution  to follow for blood grouping

  • Make sure your read the entire procedure provided by the manufacturer
  • Remember to always wear personal protective equipment such as gloves while running the test.
  • Make sure the slide is clean and dry prior to use.
  • Avoid touching antisera reagent dropper with blood sample
  • Result should be interpreted immediately after mixing.
  • Also avoid intermixing of the antisera reagents while performing the experiment as it may give false result and contamination
Also read  Difference Between Serum And Plasma

Blood grouping Procedure

  • Inform the patient or individual about the procedure to be carry out
  • Dangle the hand down to increase the flow of blood in the fingers.
  • Clean the fingertip to be pierced with spirit or 70% alcohol (usually ring or middle finger) and gently massage the finger to increase blood flow
  • With the help of the sterile lancet or pricker, pierce the fingertip and place one drop of blood in each of the four cavities
  • Now add one drop of each antiserum into each cavity respectively
ABO and Rhesus Blood Grouping Tiles
ABO and Rhesus Blood Grouping Tiles
  • Mix each blood drop with the antiserum using a fresh mixing stick or applicator stick
  • Now you can observe agglutination in the form of fine red granules within 30 seconds. Anti RhD takes slightly longer time to agglutinate compared to Anti A and Anti B.  

Blood grouping observation and Result

Blood Grouping Result and Interpretation
Blood Grouping Result and Interpretation
Slide NumberAnti AAnti BAnti RhDBlood Group
Slide 1 XA +ve
Slide 2 XB +ve  
Slide 3 AB +ve
Slide 4 XXO +ve  

Key :

  • ✓: Agglutination
  • X : No agglutination  

Blood grouping result interpretation

  • If  agglutination is observed when individual’s  blood is mixed with Anti A reagent, then the individual is said to have a blood group “A”.  
  • If agglutination is observed when individual’s  blood is mixed with Anti B reagent, then the individual is said to have a blood group “B”
  • If agglutination is observed when individual’s  blood is mixed with Anti A and Anti B reagent, then the individual is said to have a  blood group “AB”
  • If no agglutination is observed when individual’s blood is mixed with Anti A and Anti B reagent, then the individual is said to have a blood group “O”  
  • If agglutination is observed when individual’s blood is mixed with Anti RhD reagent, then the individual is said to have a “+ve” Rh factor.
  • If no agglutination is observed when individual’s blood is mixed with Anti RhD reagent, then the individual is said to a have “-ve” Rh factor.
Also read  Hemolytic Disease of the Newborn (Causes and investigation)

Common setback during Blood grouping

ProblemPossible Cause Solution
False positive result  Occurs when the antisera reagents mix with each otherOccurs when Incubated for a longer time
Being ensuring that the antisera reagents are added properly onto the respective cavity without spilling to the sides The results should be read within the time period mentioned in the manufacturer’s guide
No agglutination observed  Mostly occurs when the antisera are not stored under proper conditions Always ensure that the antisera are stored in refrigerator (2-8oC)

Source : Himelab

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About the Author: Arthur Westmann

DEFFE ARTHUR (AMOEBAMANN) is the founder and author of MLTGEEKS and MLTEXPO.He’s from Cameroon and is currently a Final year State Medical Laboratory Technician (MLT MA). Beyond lab works, he’s a passionate internet user with a keen interest in web design and blogging. Furthermore He likes traveling, hanging around with friends and social networking to do in his spare time.

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