Blood Crossmatching

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Crossmatching assesses the effect of recipient serum antibodies on donor cells (major crossmatch) and the effect of donor serum antibodies on recipient blood cells (minor crossmatch).

It is important to note that a compatible crossmatch does not completely eliminate the possibility of a transfusion reaction. Low antibody titre may not be detected via a cross match procedure.


Numerous crossmatching methods have been described and vary in their ease/speed and sensitivity. An example protocol suitable for the clinical setting is as follows:

  • Take 2ml of donor blood and 2ml of recipient blood, each into EDTA +/- serum (not a gel tube) The option exists to submit these to the laboratory for crossmatching
  • Centrifuge for 5-10 minutes (usually at around 3000rpm) and harvest the plasma. Transfer 1 drop of concentrated cells to 3ml of isotonic saline and re-suspend
  • Wash the cells in each sample 3 x in isotonic saline, filling the tubes with saline, mixing, centrifuging, and decanting
  • Finally re-suspend the cells in 3ml isotonic saline giving approximately a 2.5-5% suspension (‘weak’ tomato juice)
  • Into two tubes (or microtitre wells) mix:
Tube 1 (major


Donor cells (in isotonic saline) and recipient plasma in equal

volumes (2 drops of each)

Tube 2 (minor


Recipient cells (in isotonic saline) and donor plasma in equal

volumes (2 drops of each).

  • Incubate both tubes at 37°C for 1 hour. This incubation period can be reduced in an emergency situation, however, this will reduce the sensitivity of the test
  • Examine the tubes for any visible haemolysis or agglutination. Additionally, place one drop of fluid from each tube onto a microscope slide and examine microscopically for agglutination


  • The end point for a crossmatching is considered to be visible haemagglutination, if this is observed, an incompatibility exists

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