(2) Techniques considered optimal for hemolyzing and agglutinating
antibodies include saline or serum suspensions of donor cells incubated with recipient
serum at room temperature (18C to 25C) for 15 to 30 minutes. Although the detection
of some agglutinating antibodies is improved by incubation at 4C, any advantage
gained is outweighed by the finding of ubiquitous cold agglutinins, usually not significant
in transfusion therapy. Hence, 4C incubation is not recommended for crossmatching.
(3) The optimal method for detecting coating antibodies is the use of an
antiglobulin test following incubation at 37C for 15 to 60 minutes. Incubation may be
carried out in a potentiating medium of high dielectric constant, such as albumin. The
test is customarily read both before washing and after addition of antiglobulin serum.
b. Technical Factors. Technical factors must be considered in the
performance of a crossmatch. These include:
(1) Donor RBCs, for crossmatching, must be obtained from a sealed
segment of tubing integral with the container.
The cells used for crossmatching may be saline-washed.
A 2 to 5 percent cell suspension is usually recommended.
Reaction tubes are generally 10 or 12 X 75 mm.
(5) The supernatant must be examined for hemolysis against a white
background, before resuspending the centrifuged cells.
(6) An optical aid, such as magnifying lens, mirror, or microscope, is
advised, but not necessary, for the reading of agglutination.
(7) Hemolysis, or agglutination, at any stage of the crossmatch indicates an
(8) The person performing the test should be familiar with incubation,
centrifugation, antiglobulin technique, sources of error, reading of hemolysis, and
(9) All test tubes should be labeled before use with unit and recipient
Two accepted procedures are given below; other techniques may also be