Affinity tuning of antibodies using yeast surface display
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Affinity tuning of antibodies using yeast surface display. Yeast surface display of I-Ani1: Two epitope tags, One “antigen” binding site. Yeast surface display of I-Ani1: single bp resolution of binding specificity. 1 bp change. 2 bp change. 3 bp change. 1 bp change. 2 bp change.

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Affinity tuning of antibodies using yeast surface display

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Affinity tuning of antibodies using yeast surface display

Affinity tuning of antibodies using yeast surface display


Affinity tuning of antibodies using yeast surface display

Yeast surface display of I-Ani1: Two epitope tags, One “antigen” binding site


Affinity tuning of antibodies using yeast surface display

Yeast surface display of I-Ani1: single bp resolution of binding specificity

1 bp change

2 bp change

3 bp change

1 bp change

2 bp change


Affinity tuning of antibodies using yeast surface display

Strategies for generating variants & libraries

Rational & Computational Design

Combinatorial Libraries & Gene Shuffling

Error Prone PCR

Jim, Summer, Justin, Audrey, Umut , and others?


Affinity tuning of antibodies using yeast surface display

Optimal specific S/N ratio at physiological osmolarity

75 mM KCl

100 mM KCl

125 mM KCl

150 mM KCl

175 mM KCl

200 mM KCl

222 mOsm

262 mOsm

316 mOsm

341 mOsm

405 mOsm

472 mOsm


Affinity tuning of antibodies using yeast surface display

Analysis of computationally designed I-Ani1 variant: K24N T29R E31L R72S


Affinity tuning of antibodies using yeast surface display

JH&ST-Ani combinatorial Lib1.0

-ve MACS Fraction

+ve MACS Fraction


Affinity tuning of antibodies using yeast surface display

Targeting STS3 and STS4 to generate a high-affinity I-Ani1 variant


Affinity tuning of antibodies using yeast surface display

Yeast surface display of I-Ani1: ionic effects on HE specificity

ReoAni

STS3/4 Lib1.0.A2

Kd = 65nM

Kd = 20 nM

Kd = 8 nM

Standard conditions: 150mM KCl, 10mM NaCl, 5mM CaCl2, 10mM HEPES, 0.2% BSA, 0.1% Galactose, pH 7.5


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