and 37C tests separately and in duplicate with and without the addition of albumin.
The room temperature test will be strongly positive, but the 37C tests using saline-
suspended RBCs will usually be negative if handled correctly as the antibodies usually
only react up to 32C. If the 37C tests are allowed to cool, even a little, agglutination
may not occur, but complement may be bound and give a positive indirect antiglobulin
test (even one molecule of. the lgM anti-l can bind several hundred molecules of
complement). If enough time is available, tests can be allowed to settle for 1 to 2 hours
at 37C and read for agglutination before centrifugation where cooling may occur,
leading to false-positive results. The cells are then washed, as previously described, in
37C to 40C saline at 37C. Once the cells are washed free of serum, they can be
handled in the normal way and they no longer need to be kept at 37C.
(c) It should be noted that albumin, as well as enzymes, increase the
strength of the cold agglutinin reaction considerably, and great care must be taken that
the albumin is thoroughly warmed, before it is added to any of the tests. In many cases,
one has to rely on the saline system, without the addition of albumin, in order to issue
b. Paroxysmal Cold Hemoglobinuria. Paroxysmal cold hemoglobinuria (PCA)
is the rarest form of autoimmune hemolytic anemia. In the past, it was characteristically
associated with syphilis, but this association seems rare nowadays. More commonly, it
presents as an acute transitory form secondary to viral-like illnesses, particularly in
young children. It can also occur in older people as an idiopathic chronic disease.
(1) Direct antiglobulin test. The same principles apply here as in cold-
agglutinin syndrome in that the autoantibody, even though lgG, is a "cold" antibody and
sensitizes RBCs in the peripheral circulation, binding complement to the cell membrane
and then eluting from the cell into the serum at 37C. Thus, the direct antiglobulin test,
if positive, is a result of sensitization with complement components only (C3d and C4d).
(2) Eluate. As in cold agglutinin syndrome, there is usually no point in
making eluates from these complement-coated cells.
(3) Serum. The autoantibody has been shown to be an lgG antibody and is
classically described as biphasic in that it will sensitize cells in the cold and then
hemolyze them when the mixture is moved to 37C. This is the basis of the diagnostic
test for this disease, the Donath-Landsteiner test. The antibody will often agglutinate
normal cells at 4C, but only to low titers (for example, less than 1/64). As discussed
before, it will hemolyze cells If they are sensitized in the cold (for example, melting ice)
and then moved to 37C. The antibody usually does not sensitize cells "in vitro" above
(4) Specificity. The antibody has been shown to usually have specificity
within the P blood-group system (for example, anti-P). That is to say, it will react with all
cells, except the rare p, or Pk phenotype. Rare exceptions having anti-HI specificity
have been described.