Determining the Role of Vitronectin in Bacterial Resistance to Bactericidal Activity of Human Serum

To perform the analysis of vitronectin-dependent resistance to the bactericidal activity of human serum, first, culture and pellet the bacteria as described in the text protocol. Following centrifugation, resuspend the bacterial pellet with 1 volume of dextrose gelatin veronal buffer. Now add 1,500 CFU of bacteria to 100 microliters of DGVB++ containing 5% serum.

Incubate the sample at 37 degrees Celsius for 15 minutes with shaking at 300 RPM. Remove a 10-microliter aliquot from the reaction mixture at 0 minutes and 15 minutes. Place the aliquot on chocolate agar, and incubate the plate at 37 degrees Celsius overnight. After incubation of the chocolate agar plates, count the colonies appearing on the plate, and calculate the percentage of bacteria killed as described in the text protocol.