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Abstract: A dual reporter cell assay (DRCA) that allows real-time detection of herpes simplex virus (HSV) infection was developed. This was achieved by stable transfection of cells with an expression cassette that contains the dual reporter genes, secreted alkaline phosphatase (SEAP) and enhanced green fluorescent protein (EGFP), under the control of an HSV early gene promoter. Baby hamster kidney (BHK) and Chinese hamster ovary (CHO) cell lines were used as parental cell lines because the former is permissive for both HSV serotypes, HSV-1 and HSV-2, whereas the latter is susceptible to infection only by HSV-2. The DRCA permitted differential detection of HSV-1 and HSV-2 by observation of EGFP-positive cells, as substantiated by screening a total of 35 samples. The BHK-based cell line is sensitive to a viral titer as low as a single plaque-forming unit with a robust assay window as measured by a chemiluminescent assay. Evaluations of the DRCA with representative acyclovir-sensitive and acyclovir-resistant HSV strains demonstrated that their drug susceptibilities were accurately determined by a 48-h format. In summary, this novel DRCA is a useful means for serotyping of HSV in real time as well as a rapid screening method for determining anti-HSV susceptibilities. [Copyright &y& Elsevier]

Current sample processing (SP) methods for tacrolimus (FK506) immunoassays are mainly based on extraction of drug by organic solvent and divalent metal ions. Although these methods are effective for drug extraction and interference elimination, they suffer from drawbacks including inconvenience for operation, difficulties for automation and potential measurement bias. To overcome these limitations, this study describes a new SP reagent for blood cell lysis and protein denaturation. A TRFIA (time-resolved fluorescence immunoassay) was developed by using this SP reagent for whole blood FK506 quantification. Results show that blood samples could be turned into homogeneous solution after being treated by this SP reagent, and so could be directly applied to immunoassays without centrifugation. The analytical sensitivity of the FK506-TRFIA was 0.57 ng/mL, the within-run and between-run coefficient of variations (CVs) were both less than 10%. The FK506 values of 126 samples obtained by FK506-TRFIA correlated excellently with that obtained by ABBOTT FK506-CMIA (R2 = 0.982). Comparison studies also show that the FK506-TRFIA was highly resistant to endogenous interferences. These results suggest that the present SP method is a more promising chose for FK506 immunoassay, and in the meantime, its simplicity makes the whole-process immunoassay automation more feasible by obviating the necessary for centrifugation. • A new sample processing (SP) method is presented for effective drug release in tacrolimus immunoassay. • The FK506-TRFIA based on the proposed SP shows excellent performance regarding to accuracy, sensitivity and precision. • The proposed SP is unnecessary for performing centrifugation or using the volatile and flammable organic solvent. • Besides tacrolimus, the present SP method is potentially applicable to immunoassays of other small molecule analytes. [ABSTRACT FROM AUTHOR]