DIRECT IMMUNOASSAY FOR DETECTING ESCHERICHIA-COLI COLONIES THAT CONTAIN POLYPEPTIDES ENCODED BY CLONED DNA SEGMENTS

Abstract

We describe a simple in situ immunoassay for screening Escherichia coli colonies, to detect those that express polypeptide antigens encoded by cloned DNA segments. In the colony immunoassay that we have developed, only one antibody molecule need bind to the polypeptide molecule. Hence short fragments of coding sequences should be detectable, and problems imposed by the conformation of polypeptides "fused" to protein fragments encoded by the vector should be minimized. The method should allow the use of monoclonal antibodies, which bind to only one determinant. Colonies grown on nitrocellulose filters were lysed in situ and proteins from the lysates were blotted onto CNBr-activated filter pap..