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oai:www.ideals.illinois.edu:2142/82476 2023-07-11 col_2142_5131 col_2142_11615 com_2142_5130 com_2142_9130 com_2142_8903

Wittrup, K. Dane VanAntwerp, Jennifer Jewett 2015-09-25T20:44:16Z 2015-09-25T20:44:16Z 10000-01-01 1999 1999 Different methodologies were explored for improving YSD affinity maturation of anti-protein antibodies, using scFv D1.3. Using equilibrium sorted YSD with the D1.3/M3 antibodies, a single-pass enrichment factor of 125-fold (+/-65-fold) was achieved, indicating excellent differentiation between clones of only slightly different affinity. Optimal equilibrium affinity screening of a randomly mutated D1.3 library was performed with and without an osmotic stressor present in the binding reactions. Osmotic stress yielded a more diverse set of mutant clones from the screen, and recombination of selected mutations using site-directed mutagenesis produced a four-fold higher affinity mutant (MEC1). MEC1 was randomly mutagenized and screened by kinetic selection, identifying a large number of improved mutants. In both screens, library size was approximately 5 x 106, and mutagenesis was by error-prone PCR of the entire scFv gene. This affinity compares favorably to the highest affinities attained previously with phage display for anti-protein antibodies, without the necessity for large libraries or site-directed saturation mutagenesis. Made available in DSpace on 2015-09-25T20:44:16Z (GMT). No. of bitstreams: 2 license.txt: 4848 bytes, checksum: 96035ab3f5e1c23cc7138a224ce498bd (MD5) 9953167.pdf: 7255142 bytes, checksum: 1c2e61458038ff95a2bd0ff9e1e69748 (MD5) Previous issue date: 1999 Embargo set by: Seth Robbins for item 83757 Lift date: Forever Reason: Restricted to the U of I community idenfinitely during batch ingest of legacy ETDs Restricted to the U of I community idenfinitely during batch ingest of legacy ETDs U of I Only 165 p. Thesis (Ph.D.)--University of Illinois at Urbana-Champaign, 1999. http://hdl.handle.net/2142/82476 (MiAaPQ)AAI9953167 eng Biology, Molecular Affinity Maturation of the D1.3 Antibody Using Yeast Surface Display and Flow Cytometry text Chemical Engineering Chemical Engineering University of Illinois at Urbana-Champaign Dissertation Ph.D.