A flow cytometry-based immuno-titration assay for rapid and accurate titer determination of modified vaccinia Ankara virus vectors.

摘要: A flow cytometry-based immuno-titration titer assay was established to determine infectious unit (IU) and transducing (TU) of modified vaccinia Ankara (MVA) virus vectors. This titration method enumerates infected cells by measuring the expression viral protein for IU transgene TU in individual after staining with fluorophore-conjugated antibodies. It presents many advantages over standard approaches, such as TCID(50) or plaque assay, its convenience, rapidity accuracy illustrated excellent linearity reproducibility. Importantly, assays generated similar batch-specific values when testing varied MVA-derived preparations. Assay development revealed that post-infection time at which is evaluated, host cell type, blocking formation release progeny virion nocodazole, an anti-microtubule agent rifampin, a specific assembly inhibitor, are critical parameters precision, robustness, determination. An added advantage this it enables concurrent determination units product recombinant viruses. The latter demonstrated using MVA vector carrying human HER-2 gene fragment model. Hence, very versatile can be used well multiple titers simultaneously. Furthermore, readily adapted other poxvirus