Degree Type

Dissertation

Date of Award

1986

Degree Name

Doctor of Philosophy

Department

Microbiology

First Advisor

Paul A. Hartman

Abstract

Coagglutination tests with polyclonal antibodies were developed for the detection of Escherichia coli [beta]-galactosidase (GAL), [beta]-glucuronidase (GUD), and glutamate decarboxylase (GAD) in cell lysates. All three enzymes were detected in 93% of the E. coli tested; two of the three enzymes were detected in the remaining 7%. Among 42 non-E. coli, two (5%) were agglutinated by all three anti-enzyme conjugates, two (5%) were agglutinated by two conjugates, six (14%) were agglutinated by a single conjugate, and 32 (76%) were not agglutinated by any of the conjugates;Enzyme-capture assays with polyclonal antibodies detected the presence of GAL in seven of eight and GUD in all eight E. coli tested. Some GAL-positive Citrobacter freundii and Enterobacter cloacae also were positive by the enzyme-capture assay, indicating that the antibodies were not specific for E. coli GAL. Five other GAL-positive non-E. coli were negative by the enzyme-capture assay;The coagglutination tests and the enzyme-capture assay were rapid methods for the detection of GAL, GUD, and GAD in cell lysates;Polyclonal antibodies to heat-treated E. coli differed in the numbers of E. coli and non-E. coli detected by enzyme immunoassay. Antiserum to one E. coli strain reacted with all of the bacteria tested while antiserum to a second strain was positive for just two of five E. coli tested, Enterobacter agglomerans and Klebsiella ozanae. Attempts to remove the cross-reacting antibodies by adsorption were unsuccessful;Monoclonal antibodies to heat-treated E. coli varied in specificity. Antibodies from hybridoma 6H2 reacted with 35 of 68 (51%) E. coli; of the 13 non-E. coli tested, Enterobacter agglomerans was weakly positive and the others were negative. Antibodies from hybridoma 9B12 reacted with all six E. coli tested and with Enterobacter cloacae; however, 9B12 stopped producing E. coli-specific antibody. The antibodies from five hybridomas produced antibodies which reacted with a majority of the bacteria tested. Additional monoclonal antibodies that can supplement antibodies from 6H2 are needed.

DOI

https://doi.org/10.31274/rtd-180813-11777

Publisher

Digital Repository @ Iowa State University, http://lib.dr.iastate.edu/

Copyright Owner

Charles William Kaspar

Language

en

Proquest ID

AAI8825473

File Format

application/pdf

File Size

132 pages

Included in

Microbiology Commons

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