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ANTIBODY-BASED DETECTION AND QUANTIFICATION OF PECTOBACTERIUM CAROTOVORUM SSP. CAROTOVORUM

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Title: ANTIBODY-BASED DETECTION AND QUANTIFICATION OF PECTOBACTERIUM CAROTOVORUM SSP. CAROTOVORUM
Author: Bassoriello, Melissa Maria Ivana
Department: Department of Environmental Biology
Program: Environmental Biology
Advisor: Blom, Theo J.Hall, J. Christopher
Abstract: Pectobacterium carotovorum ssp. carotovorum (Pcc) is implicated in the destruction of ornamental plants in greenhouse recirculating systems. PCR-based detection and quantification of Pcc requires expensive instrumentation and knowledgeable users. This thesis describes the production of polyclonal antibodies and a single-domain antibody fragment (VHH) against Pcc lipopolysaccharide (LPS), and the development of user- friendly diagnostic assays for detection and quantification of the pathogen. Polyclonal ELISAs against heat-killed (HK) Pcc (limit of detection (LOD) = 81 CFU/ml; limit of quantitation (LOQ) = 216 CFU/ml) and Pcc LPS (LOD = 23 ng/ml; LOQ = 76 ng/ml) were developed. A preliminary user-friendly dipstick assay was also developed (≥ 105 CFU/ml). A phage display library was constructed (6.0 x 105 clones/ml), yielding one unique anti-Pcc LPS VHH. Using the Pcc LPS-specific VHH to produce affordable, user- friendly diagnostic assays is feasible since antibody fragments can be produced on a large scale through expression in Escherichia coli or Piccia pastoris.
URI: http://hdl.handle.net/10214/2314
Date: 2010-10-28


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