Crosslinking of Whole Cells

Reference:

Palva, E.T. and Randall, L.L. (1976). J. of Bacteriol.

Method:

1. Concentrate logarithmic phase cells 100-fold to give approximately 1 x 10¹⁰ cells/ml in a volume of 1 ml. After centrifugation, outer membrane permeability should not be altered (we have tested this using a microtitre nitrocefin assay) and the cells should be motile, suggesting no appreciable disturbance of the cell surface.
2. The cells are crosslinked by the addition of 0.01 – 0.1mg/ml of crosslinking reagent, reacted for 2 minutes and the reaction is terminated with excess 1M Tris-HCl, pH 8.5, as described for purified proteins.
3. Outer membranes are isolated and analysed by two-dimensional gel electrophoresis.