1 zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA JOURNAL OF r M&OBBLOGlCAL zyxwvutsrqponm ELSEV[ER Journal of Microbiological Methods 21 (1995) 61-66 A novel rapid procedure for the isolation of outer rnembrane proteins from zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQ Campylobacter jejuni Orla M.J. FIynn, Ian S. Blair* David A. McDowell Food Studies Research Centre, University of Ulster at Jordanstown, Shore Road. Newtownabbey, Co. Antrim, N. Ireland BT37 OQB, UK Accepted May 1994 zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQ Abstract A new molecular filtration-based method for the recovery and fractionation of cell envelope fragments from Cumpylobacter jejuni has been developed. The process, which uses a novel combination of filtration and selective solubilisation, offers major advantages over currently available methods. Inner and outer membranes associated with cell envelope fragments of Cumpy lobacter jejuni, recovered onto a regenerated cellulose filter under 1 bar negative pressure, can be sequentially treated with Triton X-100 and Triton X-lOO/EIDTA to yield a fraction principally composed of solubilised outer membrane protein (OMP). The method is rapid, efficient and uses low cost easily available equipment to produclz electrophoretic patterns and protein yields similar to the standard procedures used by previous workers. Keywords: Campylobacter jejuni; Isolation; Outer membrane protein 1. Introduction Membrane proteins are important determinants of pathogenicity in many bacteria [l] and several authors have previously conducted studies into the isolation of outer membrane proteins (OMPs) of Cumpy lobacter jejuni [2-51, a major cause of human gastroenteritis [6]. The two most widely accepted isolation methods are those of Blaser [2] and Logan and Trust [3]. These methods produce insoluble outer membrane fragments following selective solubilisation of the inner membrane by sarkosyl [7]. They involve the use of high-speed ultracentrifugation, at 100,000 x g for 5 h or 43,500 x g for 2.5 h respectively. * Corresponding author. Tel.: + 44 (232) 365131 Ext. 2137. Fax: + 44 (232) 362811. 0167-7012/95/$09.50 0 1995 Elsevier Science B.V. All rights reserved SSDI 0167-7012(94)00034-5