Efficacy of murine monoclonal antibodies in RBC phenotyping of DAT-positive samples

Determining the phenotype of patient RBCs that are positive by the DAT may prove problematic. Antigen typing of RBCs coated with IgG requires direct agglutinating reagents or chemical treatment (such as chloroquine diphosphate [CDP] or citric acid) to remove sufficient IgG to permit testing with IAT-reactive reagents. The citric acid elution method is commonly used in the United States; however, antigens in the Kell system are altered to the extent that they may appear to be absent by this method. There are a limited number of direct agglutinating monoclonal antibodies available. Murine monoclonal antibodies provide an additional tool for typing RBCs with a positive DAT. Five murine monoclonal IgG antibodies (anti-K: MIMA-22, MIMA-23; anti-Kp^a: MIMA-21, MIMA-27; anti-Fy^a: MIMA-19) were used in this study. Donor RBCs with known phenotypes were sensitized in vitro with alloanti-D, alloanti-c, and alloanti-K and with 20 autoantibodies (autoanti-D [n=3], autoanti-e [n=5], autoanti-Ce/e [n=5], autoanti-e+D+E [n=1], autoanti-I [n=1], and nonspecific [n=5]) to simulate a positive in vivo DAT. The sensitized RBCs were treated with CDP to remove IgG. To determine the efficacy of the murine monoclonal antibodies when testing DAT-positive samples, both sensitized and CDP-treated RBCs were tested with these monoclonal antibodies by the IAT using anti-mouse IgG. No discrepancies were noted with the unsensitized, sensitized, or CDP-treated RBCs. An exception was noted with a potent autoanti-I, where direct agglutination of the sensitized RBCs was obtained. This study demonstrates the value of using murine monoclonal antibodies to determine the phenotype of RBCs with a positive DAT caused by autoantibodies (e.g., in autoimmune hemolytic anemia) and supports previous studies showing that RBCs sensitized in vivo can be typed without chemical manipulation. Immunohematology2006;22:161–165.