International collaborative study to evaluate a candidate reference preparation to define an appropriate specified limit of anti-D in intravenous immunoglobulin products.
BACKGROUND AND OBJECTIVES: The aim of the study was to evaluate a lyophilized intravenous immunoglobulin (IVIG) preparation containing anti-D (02/228; nominal reciprocal titre of 8) for its suitability to define the maximum limit of anti-D in IVIG products when used in a proposed reference method of direct haemagglutination of papain-treated erythrocytes, in an international collaborative study. MATERIALS AND METHODS: Twenty laboratories tested 02/228 along with a negative control IVIG preparation and four IVIG samples containing different levels of anti-D. Nineteen laboratories performed direct haemagglutination methodology using papain-treated erythrocytes; five of these laboratories and one additional laboratory performed their in-house haemagglutination methodology (all indirect antiglobulin tests). RESULTS: The mode titre of 02/228, obtained by using the proposed reference method, was 8 (62.5% of tests). However, there was wide variation in haemagglutination titres between laboratories for three of the four samples. Correcting the titres of the samples relative to those of the proposed reference preparation reduced the interlaboratory variability and increased the frequency of the mode titres in three out of four samples. The indirect antiglobulin tests also showed wide interlaboratory variability and were less sensitive than the direct method in four laboratories. Eleven of the 14 laboratories that expressed an opinion considered that the level of anti-D in 02/228 was appropriate to define a specified limit. CONCLUSIONS: Our results demonstrate the necessity of using a reference preparation to define the maximum level of anti-D in IVIG products and ensure sufficient sensitivity in haemagglutination testing methodology. On the basis of these results, members of the European Pharmacopoeia Expert Group 6B recommended revision of the appropriate monograph to include this new specification and test. The Food and Drug Administration in the USA intends to adopt the same maximal specification defined by the reference preparation and to recommend the same test for the safety of IVIG products. Preparations 02/228 and 02/226 were also established by the World Health Organization as International Reference Reagents to standardize haemagglutination testing for anti-D in normal IVIG products.