Currently available methods to titrate adenoviral vectors (AdV) in the absence of a gene reporter such as GFP, are either time-consuming or not very reproducible. A Focus-Forming Assay (FFA) for quantification of infectious AdV particles followed by automated focus counting was developed using new monoclonal antibodies (mAbs) against the human adenovirus type 5. Briefly, in this method, 96-well plates of HEK293A cells were infected with 2-fold dilutions of AdV at seeding time. Forty eight hours post-infection, the cells were fixed with methanol. The cells were then incubated with each mAb followed by a FITC conjugated anti-mouse antibody. The plates were scanned and positive cells counted using an automated fluorescence microscopy system. The results of the FFA were compared with the plaque assay and the TCID₅₀ assay. The titer of six different recombinant AdV were compared using the FFA along with a commercial kit. The results were similar, but in contrast to the commercial kit for which the stained cells are counted manually, the software automatically counts the positives cells in the FFA. The automatic counting of positive cells makes the FFA a more precise and reliable assay compared to the commercial kit for titration of AdV.

目前可用的在没有基因报告基因（如 GFP）的情况下滴定腺病毒载体 (AdV) 的方法要么耗时，要么重现性不强。使用针对人类 5 型腺病毒的新型单克隆抗体 (mAb) 开发了一种用于定量感染性 AdV 颗粒并进行自动焦点计数的焦点形成测定 (FFA)。简而言之，在该方法中，感染了 HEK293A 细胞的 96 孔板在播种时用 2 倍稀释的 AdV。感染后四十八小时，用甲醇固定细胞。然后将细胞与每种 mAb 孵育，然后与 FITC 缀合的抗小鼠抗体孵育。使用自动荧光显微镜系统扫描板并计数阳性细胞。将 FFA 的结果与斑块测定和 TCID _{50进行比较}化验。使用 FFA 和商业试剂盒比较了六种不同重组 AdV 的滴度。结果相似，但与手动计数染色细胞的商业试剂盒相比，软件会自动计数 FFA 中的阳性细胞。与用于 AdV 滴定的商业试剂盒相比，阳性细胞的自动计数使 FFA 成为更精确和可靠的检测方法。