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Using Phage Display Technology to Produce Peptides Specific for Staphylococcus aureus Type 5 and Type 8

Maosa, Steficah K
http://rave.ohiolink.edu/etdc/view?acc_num=ysu1527676968813227

Abstract Details

2018, Master of Science in Biological Sciences, Youngstown State University, Department of Biological Sciences and Chemistry.
S. aureus is an important human and animal pathogen which is responsible for a variety of community and hospital acquired infections.S. aureus causes 20,000 deaths annually in the United States which exceeds the annual deaths caused by diseases such as HIV/AIDS, influenza and viral hepatitis together. Treatments of infections caused by this bacterium are difficult due to antibiotic resistance and failure of the active and passive immunity to confer the required immunity against this pathogen. Short synthetic peptides play a major role as bioprobes for the detection of microorganisms and biological molecules. Our goal is to produce a specific phage clone to S. aureus type 5 or type 8 using phage display technology. Phage clones displaying peptides specific for S. aureus were selected in previous studies. However, high non-specific background was seen in M13 ELISAs when using S. aureus type 8 as the ligand. In this study we reduced the high background by changing the blocking buffer from BSA to nonfat dry milk and by repeating trypsinization of the ligand used in the ELISA assays. However, our negative controls were still high, indicating continued non-specific binding of antibody to the plate. To investigate specific binding in the absence of whole cells, we purified capsular polysaccharide from S. aureus type 5 using digestion of bacteria paste with proteinase K, DNAse, RNAse and treatment with sodium periodate. The presence of carbohydrate in the column fractions was determined using a red tetrazolium assay. Purity of the carbohydrate was determined by a Bradford protein assay, a phosphate test to detect teichoic acid and light absorbance to detect nucleic acid. The purified carbohydrate was used as the ligand in an ELISA assay. Phage clones NLT8.7 and MMT5.2 were tested for binding. Neither NLT8.7 nor MMT5.2 showed significant binding to the carbohydrate. In future studies we hope to repeat the ELISA assays with the new block and trypsin treated ligand, and test all the phage clones previously selected.
Diana Fagan, PhD (Advisor)
Gary Walker, PhD (Committee Member)
Jonathan Caguiat, PhD (Committee Member)
82 p.
Biochemistry; Biology; Immunology; Microbiology
Staphylococcus aureus type 5; Staphylococcus aureus type 8; Phage display technology; capsular polysaccharide

Recommended Citations

Citations

  • Maosa, S. K. (2018). Using Phage Display Technology to Produce Peptides Specific for Staphylococcus aureus Type 5 and Type 8 [Master's thesis, Youngstown State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=ysu1527676968813227

    APA Style (7th edition)

  • Maosa, Steficah. Using Phage Display Technology to Produce Peptides Specific for Staphylococcus aureus Type 5 and Type 8. 2018. Youngstown State University, Master's thesis. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=ysu1527676968813227.

    MLA Style (8th edition)

  • Maosa, Steficah. "Using Phage Display Technology to Produce Peptides Specific for Staphylococcus aureus Type 5 and Type 8." Master's thesis, Youngstown State University, 2018. http://rave.ohiolink.edu/etdc/view?acc_num=ysu1527676968813227

    Chicago Manual of Style (17th edition)

ysu1527676968813227
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This open access ETD is published by Youngstown State University and OhioLINK.