For Caco-2 assay

It has to be pointed out that prediction failures of general ADME models are often related to two major sources namely the quality of experimental data used to derive the model and the interpretation of the final model. These problems are discussed in depth by Stouch et al. (2003). Some models fail as they were built from data collected from different sources and laboratories. Although this might work for some robust standardized ADME assay, it could produce incomparable data for others. Such problems have been reported for example for Caco-2 assays from different laboratories. Even if the experimental... 

Due to the high cost of cell culture, Caco-2 assays are usually used as a follow-up to PAMPA in ADME screening [78], and as a result, the sample burden for bioanalysis is not as heavy as for some first-hne assays, such as metabolic stability. There have been a number of reports in the literature that use automated optimization and single LC-MS/MS for sample analysis for Caco-2 assay support [46,79-81]. Nevertheless, Caco-2 samples pose a unique bioanalytical challenge. Unlike plasma or microsomal samples rich in proteins that help solubilize compounds and prevent adsorptive loss, Caco-2 samples are essentially aqueous buffer samples with very little protein. As a result, compounds with low solubility and/ or adsorption problems tend to exhibit poor recoveries in the assay due to precipitation and adsorptive losses [82,83]. An effective solution to this problem is the use of organic solvent to catch compounds immediately after incubation, but prior to analysis, in order to maintain solubility and prevent adsorptive loss to container surfaces. Another approach involves the addition of some protein such as bovine serum albumin (BSA) to the assay buffer system, thus reducing compound loss/ precipitation and improving recoveries [84]. 

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