1-37. USEFUL ADDITIONAL NOTES
a. Rouleaux can be identified by microscopic examination of autologous cells in
their serum. The cells adhere to one another on their flat surfaces, giving the classic
"stack of coins" appearance. These groups tend to roll and tumble together, but
disperse with the addition of saline, while true agglutination will not. True agglutinates
are opaque, inflexible, and have an irregular outline. In assessing rouleaux formation,
knowledge of the patient's clinical diagnosis and of the protein content and proportions
in the serum is helpful. Problems resulting from rouleaux can be encountered at any
step of the procedure, but usually, they occur in high-protein methods.
b. Some serums contain antibodies that will react with sodium caprylate used as
a stabilizer in the production of some albumlns. Serum containing caprylate antibody
agglutinates containing caprylate antibody agglutinates albumin-suspended cells,
including autologous cells. Despite the autoagglutination, the direct antiglobulin test is
negative. The phenomenon is caused by a fatty-acid-dependent antibody apparently
specific for sodium caprylate. Such serums do not agglutinate identical cells when
suspended in saline or in albumin prepared without caprylate. Usually all cells and the
autologous control are affected equally.
c. Previously immunized patients who have recently been transfused can very
rapidly produce antibody against one or several antigens on the transfused cells. The
antibody then reacts with the circulating antigen-positive cells and, if activity is
sufficiently strong, may also be found in the serum. The posttransfusion direct
antiglobulin test (DAT) will be positive with a mixed-field appearance, because only the
transfused cells contain the antigen.
d. Coexistence of cold and warm antibodies may be suspected when the
antibody-screening test demonstrates activity at RT, 37C, and AGT. This can be
resolved by preparing two separate panels: one incubated-from RT to 18C, or 4C, the
other at 37C. lgM antibodies can be neutralized by the use of 2-mercaptoethanol or
dithiothreitol. This is used when the mixture of antibodies has the possibility of being
lgM and lgG.
e. Reaction strength should be graded. Variations in the strength of reactions
may indicate the following:
(1) Dosage. The antibody may be reacting more strongly with cells that are
homozygous for the antigenic determinant than with the heterozygous cells. For
example, anti-M may react more strongly with MM cells than with MN. Some antibodies
that may demonstrate a dosage effect are anti-M, -N, -JKa, -JKb, some Duffy antibodies,
and some Rh antibodies.