Quantitative determination of anti‐K (KEL1) IgG and IgG subclasses in the serum of severely alloimmunized pregnant women by ELISA

BACKGROUND: Severe cases of HDN occur after the immunization of the mother with K (KEL1) antigen. To date, the only means of evaluating the concentration of anti‐K in maternal serum is by titration with an indirect antiglobulin test (IAT). A more accurate estimation of the serum anti‐K concentration is needed. STUDY DESIGN AND METHODS: An ELISA technique was developed for the determination of the absolute concentration of anti‐K IgG and IgG subclasses in the sera of alloimmunized patients. In this technique, after absorption of anti‐K on K‐positive RBCs and subsequent elution at acid pH, the concentration of anti‐K in the eluate was measured with a sensitive and reproducible ELISA. This method was validated with monoclonal and polyclonal anti‐K. It was then used to assay the sera of eight pregnant women with anti‐K immunization, associated with early fetal anemia (Hct, 7‐17%) detected between the 20th and the 31st week of pregnancy. In addition, in most of these cases, the anemia was associated with fetal hydrops. RESULTS: The anti‐K IgG concentration measured by ELISA in the sera of the eight women varied from 1.0 to 4.1 μg per mL (mean, 2.2 μg/mL). Therefore, severe and early forms of fetal anemia can be observed with a relatively low concentration of anti‐K (as compared to the concentration of anti‐D in similar cases of fetal anemia due to anti‐D). The mean proportion of each IgG subclass of anti‐K in these sera was IgG1, 95.9 percent; IgG2, 2.4 percent; IgG3, 1.3 percent; and IgG4, 0.4 percent. CONCLUSION: A simple method for quantitative estimation of anti‐K in human serum has been developed. Low concentrations of anti‐K can cause fetal anemia relatively early in pregnancy. This method should lead to a better identification of pregnant women whose fetuses are at risk for severe fetal anemia due to anti‐K.