FLOW CYTOMETRIC QUANTIFICATION OF SURFACE-DISPLAYED RECOMBINANT RECEPTORS ON STAPHYLOCOCCI

Surface display of recombinant proteins on bacteria and phages has bec ome an important tool in bioscience. To evaluate the various host syst ems, a great need exists for quantitative methods to determine the den sities of displayed proteins and peptides on the bacteria and phage su rfaces. Here we describe how a method previously applied for quantific ation of surface proteins on mammalian cells has been adapted for quan tification of chimeric receptors surface-displayed on bacteria; in thi s study, the bacteria being recombinant staphylococci. The presented m ethod takes advantage of fluorescence-activated cell sorting (FACS) te chnology and a new type of nonfluorescent plastic beads, similar in si ze (2 mu m in diameter) to bacterial cells, and thus suitable for gene ration of calibration curves from which the number of chimeric recepto rs can be obtained. The method was used to estimate the number of anti genic sires on two types of recombinant staphylococci, both carrying h eterologous chimeric receptors, and it was found that the recombinant Staphylococcus carnosus cells carried approximately 10(4) surface-disp layed antigenic sites, while recombinant Staphylococcus xylosus expose d approximately 3 x 10(3) sites per cell. The rise of the deviced meth od for different applications is discussed.