Plaque-lift testing of expression vector lambda gt11 with gold-labeled immunoglobulins.
Colloidal gold particles were coated with affinity-purified antibodies against the human plasma protein, C1 inhibitor, and used to probe for fusion proteins of C1 inhibitor with beta-galactosidase encoded by recombinant bacteriophage lambda gt11 DNA. Plaque-lift tests were done with recombinant proteins immobilized on nitrocellulose applying anti-C1 inhibitor gold particles followed by the silver enhancement treatment. This procedure resulted in a sensitive and specific staining of the recombinant proteins and allowed the selective detection of relevant clones in a complex cDNA expression library. Under optimized conditions, plaque-lift testing was completed within 2.5 h after removal of nitrocellulose filters from the plate. Hence, the immunogold detection method provides an alternative to conventional enzyme- or radionuclide-based screening procedures for cDNA expression libraries.
Pohl
H E
HE
Department of Clinical Biochemistry, University of Munich, Federal Republic of Germany.
Hock
J
J
Müller-Esterl
W
W
D000818:Animals; D010582:Bacteriophage lambda / Q000235:genetics / Q000254:growth & development; D003001:Cloning, Molecular / Q000379:methods*; D003102:Colloids; D003174:Complement C1 Inactivator Proteins / Q000032:analysis / Q000235:genetics* / Q000276:immunology; D004247:DNA / Q000235:genetics; D005822:Genetic Vectors; D006046:Gold; D006801:Humans; D007074:Immunoglobulin G; D008297:Male; D011993:Recombinant Fusion Proteins / Q000032:analysis / Q000276:immunology; D013552:Swine; D010948:Viral Plaque Assay
D003102:Colloids; D003174:Complement C1 Inactivator Proteins; D007074:Immunoglobulin G; D011993:Recombinant Fusion Proteins; D006046:Gold; D004247:DNA