BIFICO is a main microecological drug that the main ingredient is Bifidobacterium longum ATCC BAA- 2753. It’s necessary to detect the number of Bifidobacterium longum ATCC BAA-2753 for quality control during the production of BIFICO. Plate count assay (PCA) is the most commonly used method for counting microbial cells. However, not all microorganisms can be counted by PCA. (e.g. uncultured microorganisms under specific growth conditions). In this study, a method of fluorescent techniques in combination with flow cytometry (FCM) was established for the counting of Bifidobacterium longum BAA-2753. Using SYTO9 and PI stain, flow cytometric analysis could easily differentiate live bacteria and dead bacteria. The number of live bacteria determined by FCM was about fourfold higher than that counted by PCA. Therefore, the FCM method could be a practical tool for the quality control of Bifidobacterium longum BAA-2753 in the industrial production of BIFICO.