Biological and immunological characterization of human luteinizing hormone: IV. Biological and immunological profile of two international reference preparations after electrofocusing

Abstract

The first IRP of Human Pituitary Gonadotrophins (FSH/LH) for bioassay ( 69 104 ) and the 1st IRP of Human Pituitary Luteinizing Hormone (LH) for immunoassay ( 68 40 ) were fractionated by an electrofocusing technique in a sucrose density gradient and the profile of biological and immunological activities was determined. The partially purified LH preparation ( 69 104 ) gave a broad pattern of biological and immunological activities which extended from pH 4 to 10. The biological : immunological (B/I) ratio of the various fractions (using the 68 40 preparation as a standard) ranged from 0.04 to 1.05. The low B/I ratios indicate the presence of high levels of immunologically active, biologically inactive material in this preparation. In contrast to the behaviour of the 69 104 preparation, the major proportion (88%) of the biological activity recovered after electrofocusing of the highly purified LH preparation ( 68 40 ) was found within the pH range 7–9, with B/I ratios (again using as standard the 68 40 preparation) ranging from 0.4 to 1.5. Multiple dose parallel line design radioimmunoassays revealed a lack of parallelism between the 69 104 and 68 40 International Reference Preparations. This was attributed to the presence of acidic material in the former preparation, which is practically absent from the latter. The biological LH profiles of both the 69 104 and 68 40 preparations differed from those previously reported for aqueous extracts of pituitaries. It is concluded that the dissimilarity in the electrofocusing profiles of the biological and immunological activities of the 69 104 preparation renders this preparation unsuitable as a reference preparation for the quantitation of biologically active LH by radioimmunoassay methods. Using the same criteria, the 68 40 preparation would appear to be a more suitable standard.