Reducing the variation in performance of antibody titrations

Antibody titration is difficult to standardize. We investigated whether a detailed, uniform procedure for antibody titration would reduce variation in both tube-based and gel card titres in an international study.

Laboratories (n = 35) tested proficiency testing material provided by the College of American Pathologists each according to (i) their routine method; (ii) a detailed, uniform method; and (iii) the uniform method titrating the serum sample against a red cell of specified phenotype (D+ C- c+ E+ e- for anti-D; A(1) for anti-A) instead of the red cell of the same phenotype provided in the proficiency testing kit. Uniform method results were reported with 1+ and w+ end-points. Paired statistical analyses of variance were conducted using the F-test.

The variance between laboratories was not significantly reduced with the uniform method using a 1+ end-point. However, a statistically significant reduction in the variance of anti-D and anti-A titres by the tube-based uniform technique after 37 degrees C incubation and conversion to the antiglobulin (AHG) phase was seen when 19 laboratories reanalyzed their results using a w+ end-point. Too few laboratories reported results with a w+ end-point in gel card testing to allow analysis. Titration against red cells of the specified phenotype provided by the participating laboratory did not appear to introduce additional variance. Overall, results reported based on the gel card technique at the AHG phase (1+ end-point) showed reduced variance compared to tube-based techniques.

A detailed, uniform method for antibody titration at 37 degrees C and read at the AHG phase in a tube-based method with a w+ end-point reduced interlaboratory variability.