Publication Date
1-1-2002
Document Type
Dissertation/Thesis
First Advisor
Mitchell, John L. A.
Degree Name
B.S. (Bachelor of Science)
Legacy Department
Department of Biological Sciences
Abstract
Antibodies are proteins of the immune system that exhibit exquisite specificity to their binding partners, called antigens. Binding occurs at the hypervariable region (Fv), which is capable of discriminating even enantiomeric molecules. By targeting on the DNA sequences that code for Fv and expressing it as a protein, the binding properties of an antibody can be assessed. In this project, the heavy and light chains of the Fv region (Vh and V[_) of an anti-amino acid antibody (anti-phenylalanine 67.36) were linked using a short nucleotide linker and four primers designed from previously determined sequences of Vh and V|_. The resulting V|-H¡nker-V|_(HIL) construct - a single chain variable fragment (67.36 HIL scFv) - was inserted into a TOPO cloning vector and transformed into E. coli BL 21 expression cells. 1 L cultures were grown at 37°, 30° and 27 °C. Cells were collected, resuspended and lysed to release the protein of interest. Isolation and purification was performed using a metal (cobalt) affinity chromatography resin. Characterization by SDS-PAGE indicated a correlation between incubation temperature and extent of expression. Noncompetitive ELISA tests revealed a functional 67.36 HIL scFv. However, further tests e.g., competitive ELISA, must be carried out to confirm an enantiomeric preference.
Recommended Citation
Lezondra, Lour-Evelyn, "Biotechnological production of a stereoselective scFv antibody fragment" (2002). Honors Capstones. 238.
https://huskiecommons.lib.niu.edu/studentengagement-honorscapstones/238
Extent
19 unnumbered pages
Language
eng
Publisher
Northern Illinois University
Rights Statement
In Copyright
Rights Statement 2
NIU theses are protected by copyright. They may be viewed from Huskie Commons for any purpose, but reproduction or distribution in any format is prohibited without the written permission of the authors.
Media Type
Text
Comments
Includes bibliographical references.