c. Specificity of the Antibodies Eluted from the Patient's Red Blood Cells
and in the Serum.
(1) Red blood cell eluate. It is essential to prepare an eluate from the RBCs
in order to define the autoantibody. If the positive direct antiglobulin test is a result of
sensitization with complement components only, no antibody is usually detectable in the
eluate, from the RBCs. If lgG is present on the cells, it can be eluted by simple
methods, (for example, heat or ether). The eluate should contain autoantibody only,
(this can often be confirmed by showing the presence of the matching antigen on the
patient's RBCs). If the direct antiglobulin test Is positive, as a result of lgG sensitization,
and the eluate shows no activity against normal cells, an association with drugs should
be strongly suspected. A good example of this is penicillin-induced hemolytic anemia,
where, the eluate from a strongly positive direct antiglobulin test will not react with any
untreated normal cells; however, it will react strongly with the same cells treated with
The serum may contain:
1 No antibodies; they may all have been autoabsorbed onto the
patient's RBCs "in vivo".
2 Autoantibodies only.
3 Autoantibodies plus alloantibodies.
4 Alloantibodies only.
(b) Comparing the specificity of antibody present in an eluate from the
RBCs with antibody in the serum helps indicate the presence of alloantibody in the
serum. The patient's RBC phenotype may help to confirm this.
(c) The specificity of the autoantibodies associated with warm antibody
AIHA is very complex. The main specificity is directed against the Rh complex, but may
only be obvious if rare cells, such as -D-, Rhnull, LW-negative, U-negative, and
Wrb-negative are available. If these cells are available, one can demonstrate "Rh"
specificity in the broadest sense, in approximately 70 percent of these cases. Clear-cut
specificity, such as anti-RhO(D), rh'(C), rh"(E), hr'(c), and hr"(e) is present only rarely,
anti-hr"(e), being the most common. Usually, the undiluted eluate or serum will react
with all RBCs of common Rh phenotype, but variations in strength of reaction may be