Single-cell detection: Test of microbial contamination using capillary electrophoresis

Single cells of bacteria and fungi were detected using a capillary electrophoresis-based test for microbial contamination in laboratory samples. This technique utilizes a dilute cationic surfactant buffer to sweep microorganisms out of the sample zone and a small plug of blocking agent to negate the cells' mobility and induce aggregation. Analysis times are generally under 10 min. Previously, a nutrient broth media was reported as an effective blocking agent; however, the natural background fluorescence from the nutrient broth limited the detection sensitivity to similar to 50 cells. In order to enhance the sensitivity of the technique down to a single cell, an alternative synthetic blocking agent was sought. Various potential blocking agents were screened including salts, polypeptides, small organic zwitterions, and surfactants. Zwitterionic surfactants are shown to be attractive alternatives to a nutrient broth blocker and mimic the nutrient broth's effects on cellular aggregation and mobility. Specifically, caprylyl sulfobetaine provided the sharpest cell peaks. By substituting caprylyl sulfobetaine in place of the nutrient broth, the fluorescence of the blocker plug was reduced by as much as 40x. This reduction in background noise enables detection of a single microorganism in a sample and allows this technique to be potentially used as a rapid sterility test. All single cells analyzed using this technique displayed signal-to-noise ratios between 5 and 9.