Purification of flagellar cores of Vibrio cholerae

A procedure is described for the purification of the cores of flagella sheared from Vibrio cholerae. V. cholerae is a monotrichous organism whose flagellar core (FC) is enclosed within a sheath. The purification procedure consists of several cycles of differential centrifugation and cesium chloride density-gradient ultracentrifugation in the presence of a neutral detergent, Triton X-100. Purity of the FC preparations is assessed by electron microscopy, polyacrylamide gel electrophoresis, and chemical analysis. The purified FC preparations are devoid of flagellar sheaths and free from detectable cell wall and cytoplasmic contamination. Antibody prepared in rabbits against purified FC reacts with the flagellum of intact V. cholerae or purified FC as seen by ferritin-labeled antibody studies. Purified FC is composed of a single protein subunit with an estimated molecular weight of 45,000 g/mol and a density of about 1.3 g/cm3.