POLYCLONAL ANTIBODIES AGAINST LIVE CELLS OF PSEUDOMONAS-FLUORESCENS FOR THE DETECTION OF PSYCHROTROPHIC BACTERIA IN MILK USING A DOUBLE ANTIBODY SANDWICH ENZYME-LINKED-IMMUNOSORBENT-ASSAY

Polyclonal antibodies raised against live cells of Pseudomonas fluores cens AH-70 have been used in a double antibody sandwich ELISA techniqu e for the detection of P. fluorescens and related psychrotrophic bacte ria in refrigerated milk. These antibodies, bound to the wells of a mi crotiter plate, were used to capture the microorganisms on milk sample s. Further immunorecognition of the captured antigens was attained wit h the same antibodies conjugated to biotin. ExtrAvidin(R)-peroxidase w as used to detect the biotinylated antibodies bound to their specific antigens. Subsequent enzymic conversion of substrate showed dear diffe rences in absorbance after assay of milk samples containing P. fluores cens strains of different origin and related psychrotrophic microorgan isms. The detection threshold for the sandwich ELISA assay developed i n this work was 10(5) cfu/ml.