KNOWLEDGE CENTRE

Typing red cells for antigens other than ABO and Rh(D) is not done for routine pretransfusion testing in which the patient lacks unexpected antibodies. However, if patients are found to have irregular antibodies, they are typed for the corresponding antigens to confirm if the antibodies are alloantibodies or autoantibodies. Also, if such a patient requires blood transfusion, all donor units to be crossmatched are antigen typed.

Note: If a crossmatch with donors was done in parallel with antibody screening, any incompatible donor units are antigen typed to confirm that they are indeed antigen positive, thus explaining their incompatibility. [Some labs may not antigen type incompatible donors due to cost constraints.]

If an incompatible donor was found to be antigen negative, then another explanation would have to be found to explain the agglutination in the crossmatch, e.g., perhaps the donor's cells have a positive DAT. Any apparently compatible donor units must also be antigen typed to be sure they really do lack the corresponding antigen and thus are safe to transfuse.

Typically, donors are antigen typed before they are crossmatched. Donor cells may be positive for the corresponding antigen and still be compatible with the patient's serum if the patient has a relatively weak antibody whose corresponding antigen exhibits dosage. In such a case, the patient's antibody may only be able to agglutinate red cells from homozygous and may be non-reactive with a donor who has the antigen only in the heterozygous state. The heterozygous form of the antigen will be able to be detected by commercial antisera, however, since it contains potent antibodies capable of agglutinating red cells with weak, heterozygous antigens.

There are three controls necessary when typing for red cell antigens:

1. Control of the antisera

A positive control which ensures the reactivity of the antisera. The positive control cell should be weakly reactive for the antigen being tested, i.e., from a heterozygous donor.
A negative control which ensures the specificity of the antisera.
Red cells for these controls are selected from among the panel used for antibody identification.

2. Control of the cells being phenotyped

An autocontrol which ensures that the cells being tested will not spontaneously agglutinate with the antisera, even if they lack the corresponding antigen. This can happen, for example, if the red cells being typed have a positive DAT and are agglutinated by the protein in the antisera. Or, if the antigen typing is to be done by an IAT test, and the test cells have a positive DAT, they will agglutinate once antiglobulin serum is added, even if they lack the antigen being typed. Due to cost constraints some labs do not do autocontrols when typing donors and do not do an autocontrol on the patient if a DAT has been done earlier as part of the antibody identification.

In general, an autocontrol should parallel the conditions of the antigen typing test, except for being exposed to the specific antibody in the antiserum. Thus autocontrols will vary according to the type of antisera being used to antigen type. For example:

For saline-reactive antisera such as saline anti-D (which contains IgM anti-D in a low protein concentration of about 6 or10%, depending on the supplier), a suitable autocontrol (Rh control) would be

1. Negative reaction with any test that includes the patient's red cells in a low protein medium, e.g., ABO typing sera. Using this protocol, only patient's who appear to be group AB Rh(D) positive need to have a control set up.
2. Patient's red cells + 6% albumin (or whatever percent protein is in the typing sera.)

Note: An autocontrol when Rh typing using monoclonal/polyclonal blend anti-D (which contains a blend of IgM and IgG anti-D in a low protein concentration) would be similar to that for saline-reactive antisera as described above.

For "slide and modified tube" antisera (containing an IgG antibody in a high protein concentration of about 22% - 30%), a suitable autocontrol would be patient's red cells + manufacturer's suppled control (i.e., the Rh control medium supplied with high protein anti-D)

For "IAT only" antisera (containing an IgG antibody which can only be detected by doing an IAT test and adding antiglobulin serum), the autocontrol would be Direct antiglobulin test (DAT), i.e., wash patient's red cells 4 times and add antiglobulin serum. A negative DAT confirms that antigen-negative red cells will not agglutinate with antiglobulin serum.