Assay strategies comparability testing

For a majority of biomarker assays, standard calibrators are prepared in an analyte-free alternative matrix instead of the de facto sample matrix of patient samples. For such methods, it is crucial to demonstrate that the concentration response relationship in the sample matrix is similar to that of the alternate matrix. Spike-recovery experiments with the reference standard may be inadequate to evaluate the matrix effect, as the reference standard may not fully represent the endogenous analyte. Instead, parallelism experiments are performed through serial dilutions of a high-concentration sample with the calibrator matrix. Multiple individual matrix lots (>3 lots) should be tested to compare lot-to-lot consistency. In the instance that the limited amounts of sample are available, apooled matrix strategy can be used with caution as discussed by Lee et al. [15]. The samples can be serially diluted with the standard matrix (standard... 

Two main fields of steroid analysis described in various pharmacopoeias are worthy of mention the identification and purity testing of steroid raw materials and formulated products. The USP also recommends TLC for steroid assay, but the number of these applications is decreasing from edition to edition, as TLC methods based on spot elution are replaced by HPLC methods. It should be noted that different strategies (performance of TLC procedures, evaluation of the separated spots) are described by the various pharmacopoeias the TLC methods described for the analysis of steroid raw materials and formulations in USP XXII (28, Table 4), BP88 (29, Table 5), and Hi.Eur. (37, Table 6) are compared in Table 7. 

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