The fourth International Standard for Human Urinary FSH and LH: specificities of LH seminal vesicle weight gain assays in the collaborative study differ between laboratories

Abstract

The fourth International Standard for Human Urinary FSH and LH (IS; in ampoules coded 98/704) was compared with the third International Standard for Urinary FSH and LH (IS 71/264) by 10 laboratories in nine countries, using FSH and LH in vivo bioassays.Estimates of the FSH content of the IS by augmented ovarian weight gain assays were homogeneous within each laboratory and over all laboratories. The combined weighted geometric mean estimate of FSH content of the IS (with 95% fiducial limits) in terms of IS 71/264 was 71·9 (69·0-74·9) IU/ampoule.Although estimates by seminal vesicle weight gain (SVW) assays of the relative LH activities of the IS and IS 71/264 were homogeneous within laboratories, estimates were heterogeneous between laboratories. This indicated differences between the spectrum of LH isoforms in the IS and IS 71/264, which were obtained from different manufacturers, and differences between the specificities of SVW assays performed in different laboratories. The differences between the specificities of SVW assays appeared to be related to interactions among mean laboratory seminal vesicle weights, age and genetic strain of rat. The finding of inter-laboratory differences in the specificities of SVW assays is of some significance, as this assay method has been generally adopted by Pharmacopoeias for the control of the LH content of therapeutic products. The combined unweighted geometric mean estimate of LH content of the IS (with 95% fiducial limits) in terms of IS 71/264 by SVW and ovarian ascorbate depletion assays was 70·2 (61·7-80·0) IU/ampoule.Estimates of the FSH and LH content of ampoules of the IS kept at increased temperatures suggested that the IS would be adequately stable under normal storage conditions.On the basis of these results, the World Health Organization Expert Committee on Biological Standardization established the preparation in ampoules coded 98/704 as the fourth International Standard for Human Urinary FSH and LH, and assigned to the contents of each ampoule an activity of 72 International Units of urinary FSH and an activity of 70 International Units of urinary LH.

Role of Ancillary Variables in the Design, Analysis, and Interpretation of Animal Experiments

“…Analysis of ancillary variables may nevertheless provide indications of variables that could be relevant in future research. For example, the study by Storring and Gaines Das (2001) suggested that the exact age of the rats used in those experiments may be a more critical variable than had previously been recognized, and that this factor may also be strain dependent to some extent, even though these questions were not directly addressed by that study.…”

“…In a recent study (Storring and Gaines Das 2001) in which ovarian weight relative to body weight was measured in rats as response to follicle stimulating hormone, body weights and organ weights were also reported separately for many experiments. A marked outlying response was detected in one experiment in which weights of the 48 female rats were within the range 73 to 90 g with the exception of one, which was reported as 58 g. When the data were analyzed, it could no longer be determined whether this was a recording error; however, the difference was so extreme that it was omitted from analysis.…”

“…Such failure is unethical if it leads to unnecessary repetitions of an experiment. A recent international study (Storring and Gaines Das 2001) revealed the extent to which ancillary variables can assist in accounting for between laboratory differences. This study used an assay method adopted by pharmacopoeias for the control of therapeutic products, and hence a method that might be considered to be well defined.…”

Assessment of the biopotency of follitropin alfa and lutropin alfa combined in one injection: a comparative trial in Sprague-Dawley rats

“…The National Institute of Biological Standards and Controls (NIBSC) Urinary Follicle Stimulating Hormone and Urinary Luteinizing Hormone, 4 th International Standard, NIBSC Code: 98/704 (NIBSC, Herts, UK) reference standard was prepared for each replicate assay by dissolving a single vial (FSH assay) or two vials (LH assay) each with 1.0 ml sterile saline solution and subsequently pooled, in the case of the latter. By definition of the World Health Organization Expert Committee on Biological Standardization, the NIBSC 98/704 standard contains 72 IU of urinary FSH and 70 IU of urinary LH per ampoule [ 18 ]. The NIBSC 98/704 standard was used because a similar standard was not available for a combination of recombinant FSH and recombinant LH.…”

Análise de luteotrofina humana e de gonadotrofina coriônica humana, recombinante e natural, por cromatografia líquida de alta eficiência em fase reversa

“…horas ("overnight") a 37 °C com 5 M ácido acético e aplicando o produto da reação de dissociação diretamente na RP-HPLC (Carvalho, 2009). (Storring, 2001).…”