Bioanalytical Screening Methods

High-pressure liquid chromatography (HPLC) formats that use reverse-phase analytical columns dominate the bioanalytical field. Isocratic elution formats are used for traditional LC-MS/MS bioanalysis, since these separations do not require additional time for column reequilibration. However, gradient elution is a more effective approach for cassette analysis in highthrough-put screening methods for discovery applications because of its versatility for the separation of compounds that have a wide variety of lipophilicity. 

Mazzarino, M., de la Torre X., and Botre, F. (2008) A screening method for the simultaneous detection of glucocorticoids, diuretics, stimulants, anti-oestrogens, beta-adrenergic drugs and anabolic steroids in human urine by LC-ESI-MS/MS. Analytical and Bioanalytical Chemistry, 392, 681-698. 

Pozo, O.J., Van Eenoo, P., Deventer, K., and Delbeke, F.T. (2007) Development and validation of a qualitative screening method for the detection of exogenous anabolic steroids in urine by liquid chromatography-tandem mass spectrometry. Analytical and Bioanalytical Chemistry, 389,1209-1224. 

The second group can be represented by single bead methods, and relies on either bioanalytical methods to select the active compounds or on-bead screening to determine the beads carrying active compounds. It is limited to solid-phase chemistry and does not require chemical steps after library synthesis but does require sophisticated analytical methods to determine the structure of the active compounds. A recent hybrid deconvolution-single beaddecoding method named DRED (dual recursive deconvolution) requires both deconvolutive techniques and sophisticated analytical capacities. 

A myriad of published reports has now proven the broad and multifaceted applications of modern MS-based techniques for the analysis of small molecules [89-103], Higher-throughput screening has been in demand and will continue to be one of the main objectives of industrial laboratories. However, bioanalytical scientists should bear in mind that the quality of science cannot be compromised at the expense of speed. To this end, poorly developed LC/MS-based methods that lack specificity, sensitivity, and/or ruggedness can lead to erroneous or misleading PK readouts. 

As for any bioanalytical method, the extent of validation for an immunoassay should be related to the intended application of the assay. Thus, if an immunoassay is intended to support rapid screening in discovery R D, the characterization of specificity and the accuracy and precision specifications may be less stringent than if the assay is used to support pre-clinical and clinical development studies. Indeed, an assay for discovery support may be designed to detect active metabolites as well as parent molecule, so that... 

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