c. The procedure for the direct antiglobulin test is presented below.
(1) STEP 1. Place one drop of a 2 to 5 percent saline suspension of cells to
tested in a labeled 10-x 75-mm tube. Wash 3 or 4 times with saline. After last wash,
decant completely. Add one or two drops of antiglobulin serum: mix.
(2) STEP 2. Centrifuge and examine for agglutination with an optical aid;
grade and record results. (The manner in which the RBCs are dislodged from the
bottom of the tube is critical. The tube should be held at an angle and shaken gently
until all cells are dislodged. Then it should be tilted gently back end forth until an even
suspension of cells or agglutinates is observed.)
(3) STEP 3. To control for inadvertent contamination of the antiglobulin
serum, add one drop of lgG-sensitized RBCs to any tubes that have been recorded as
negative and recentrlfuge. If the patient's cells were washed adequately in the first
stage of the test, the control cells should be agglutinated, and the negative result on the
patient is valid.
If monospecific anticomplement (-C3, -C4 or nongamma) reagents are used
(para 1-21), complement-sensitized cells should be substituted for the
lgG-sensitized cells in step 3.
1-17. INDIRECT ANT IGLOBULIN TEST
a. The indirect antiglobulin test is used to demonstrate antibodies that may
cause RBC sensitization "in vitro". The antibody-containing serum is incubated with
specific RBCs, which, following washing, are reacted with antiglobulin serum to see
whether RBC sensitization has occurred.
b. The indirect antiglobulin test is useful for:
Detection and identification of unexpected antibodies.
Detecting RBC antigens not demonstrable by other techniques.
Special studies, (for example, leukocyte and platelet antibody tests).
c. The technique of the indirect antiglobulin test is included in other sections of
this manual where this technique is utilized, (for example, Lesson I, Sections Ill and IV).