Quantification of Follicle Stimulating Hormone (follitropin alfa): Is In Vivo Bioassay Still Relevant in the Recombinant Age?

R. Driebergen, G. Baer

Disclosures

Curr Med Res Opin. 2003;19(1) 

In This Article

Limitations of the Steelman-Pohley FSH Bioassay

Although the Steelman-Pohley bioassay has proved to be a robust, specific in vivo assay for FSH, and remains the mainstay of pharmacopoeial monographs for the statutory determination of the FSH potency of therapeutic preparations, it has a number of important limitations. The most important of these are: the assay's limited precision, its requirement for the sacrifice of large numbers of animals and its cumbersome data generation and interpretation procedures, which require highly controlled conditions and standardised procedures.

The bioassay is widely acknowledged to have only limited precision - its coefficient of variation (CV) in a single determination is 10-20%, implying that the acceptable range of FSH activity in proprietary preparations is highly variable. A vial that nominally contains 75 IU of FSH on the basis of an initial assay meets the 80-125% (i.e. 60-94 IU) acceptance range and the 64-156% (i.e. 48-117 IU) fiducial limits. However, in reality the vial may contain anything from 48 IU to 117 IU FSH.[5] Furthermore, because biological systems are not entirely predictable, their use in the context of bioassays leads to variable responses between tests and between test laboratories. Bioassays therefore require highly standardised conditions. In order to reduce variability to approximately 10%, and to obtain a sufficient number of valid tests to calculate the final test result, multiple replicates, multiple standard points and multiple independent tests are required. Consequently, it is very common for 150-200 immature rats to be sacrificed to obtain a single final test result per sample. Given that, on an industrial scale, several hundreds of samples are submitted for testing each year, it is not unusual to find that tens of thousands of animals are being killed annually. These days, there is increasing ethical and political pressure to reduce the use of laboratory animals.

Overall, therefore, the Steelman-Pohley bioassay is cumbersome, time-consuming, and lacks precision. There is a clear need for a more precise, and more ethically acceptable, physico-chemical alternative to measure the potency of r-hFSH preparations.

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