International collaborative study to evaluate methods for quantification of anti-D in immunoglobulin preparations.

ABSTRACT

BACKGROUND AND OBJECTIVES: The disadvantages of autoanalyser methodology for anti-D potency estimation have prompted the search for an alternative reference method. The aim of this study was to carry out a direct comparison of autoanalyser methodology, competitive enzyme-linked immunoassay (EIA) and flow cytometry. MATERIALS AND METHODS: The anti-D potencies of nine immunoglobulin preparations were estimated against the World Health Organization (WHO) International Reference Preparation for anti-D immunoglobulin, using the three different assay methods described above, in an international collaborative study involving 18 laboratories. RESULTS: No significant differences in potency estimates for five of nine samples were identified using the three different methods. For six of nine samples the mean potency estimates obtained using competitive EIA lay between those of the autoanalyser and flow cytometry. Flow cytometry gave the lowest estimates for seven of nine samples. The range of intralaboratory variability, expressed as percentage geometric coefficients of variation (% gcv) and excluding extreme values, was as follows autoanalyser (eight laboratories), 0.4-17.0; competitive EIA (12 laboratories), 2.6-34.8; and flow cytometry (eight laboratories), 2.9-30.0. The range of interlaboratory variability (expressed as % gcv) was autoanalyser (eight laboratories, eight samples), 10.8-17.6; competitive EIA (12 laboratories, nine samples), 10.3-17.3; flow cytometry (nine laboratories, eight samples), 6.2-16.1. CONCLUSIONS: The study did not show clear superiority of one method over the others. Both competitive EIA and flow cytometry are acceptable alternatives to autoanalyser methodology for polyclonal anti-D potency estimation. For monoclonal anti-D, further work is necessary to determine the most appropriate method for potency testing.