A quantitative immunoassay utilizing Escherichia coli cells possessing surface-expressed single chain Fv molecules.
A facile, quantitative immunoassay is described that utilizes Escherichia coli (E. coli) bacteria expressing single chain Fv (scFv) antibody fragments attached to the cell surface. A Scatchard analysis demonstrated that the antibodies on the surface of the cells retained full binding activity (Kd = 2.2 x 10(-9) M) and that there are 60,000 scFv molecules per cell. The cells are used as the antibody reagent in the assay, and, following incubation with analyte, simple centrifugation is used to separate the antibody-bound from unbound analyte. The immunoassay is rapid and accurate down to the nanomolar level. In addition, a variety of detection strategies can be used, and the immunoassay is not adversely affected by the presence of animal serum. A key advantage of the new immunoassay is that the antibody reagent can be inexpensively produced in a "ready to use" form by simply growing cultures of the bacteria.
Chen
G
G
Department of Chemistry and Biochemistry, University of Texas at Austin 78712, USA.
Cloud
J
J
Georgiou
G
G
Iverson
B L
BL
D000954:Antigens, Surface / Q000276:immunology; D002460:Cell Line; D004077:Digoxin / Q000032:analysis; D004926:Escherichia coli / Q000302:isolation & purification / Q000378:metabolism*; D016231:Fluorescence Polarization Immunoassay; D007128:Immunoglobulin Fragments / Q000096:biosynthesis; D007700:Kinetics; D011863:Radioimmunoassay
D000954:Antigens, Surface; D007128:Immunoglobulin Fragments; D004077:Digoxin