Detection and sorting of microbial cells and sub-micron particles
Introduction
While most modern applications of flow cytometry may focus on cells of eukaryotic origin, the first flow analyzers were developed with bacterial detection in mind—an application that is still very much of interest to biologists.1 Unlike mammalian cells, prokaryotes are, at most, a couple microns wide and scatter very little light.2 Additionally, extracellular vesicles (EVs) that fall into a similar size category (0.03 to 1 μm) form yet another modern important scientific spotlight. Collectively, prokaryotes and EVs can be considered “small particles” for the purpose of flow cytometric analysis. The WOLF cell sorter can be utilized for a variety of such experiments. Here, we present the practical limits of detection for the WOLF, as well as examples of microbial detection and sorting data.
Method
Sensitivity testing
Size standards from Spherotech (CAT #PPS-6K) were diluted 10-fold in PBS and used to demonstrate resolution in the 2-15 μm range using the predetermined “Rainbow” setting on the WOLF (FSC threshold of 8,400). A sub-micron calibration bead mix from Apogee Flow Systems (CAT #1493) containing equivalent ratios of silica and polystyrene standards were diluted 50-fold in PBS to achieve a concentration of 1000 total events per second on the WOLF using settings of ...
Want to read more?
