Purpose-Oriented Antibody Libraries Incorporating Tailored CDR3 Sequences
Abstract
:1. Introduction
2. Results and Discussion
2.1. Long CDRH3 Libraries for the Inhibition of Enzymes
2.1.1. Characterization of Libraries by Deep Sequencing
CDRH3 source | Library | Size | Unique CDRH3 | |
---|---|---|---|---|
Number | % | |||
Human | HnB | 2.9 × 108 | 313'094 | 38.1% |
Mouse | MnA | 2.5 × 108 | 70'993 | 13.0% |
Synthetic long | SL | 5.2 × 109 | 398'124 | 43.6% |
*185'453 | *23.3% |
2.1.2. Assessment of Library Performance by Phage Display Selections against HRP
Clone | CDRH3 | CDRL3 | CDRH3 length | HRP inhibition IC50 [nM] |
---|---|---|---|---|
MnA-A3 | ARDAHGWYF----------DV | GTWDMPPD-VV | 11 | - |
MnA-F5 | ARSYANYDWF---------DY | QQSVHYRP-ST | 12 | - |
MnA-H4 | ARLGTGTGYF---------DV | GTYDVGRV-HV | 12 | - |
MnA-H11 | ARQGNGYY-----------AY | QQSEGVP--WT | 10 | - |
HnB-A4 | ARPQTYYYDSSGYPDAF--DI | QQREVLP--LT | 19 | - |
HnB-B9 | ARGWGRTAT----------DY | QQPARQWP-RT | 11 | - |
HnB-F9 | ARDPGVLWSSSSPYYF---DY | QQDLAGWP-BT | 18 | - |
HnB-G4 | ARGLVFDSSGYYGF-----DY | QQPSPMP--PT | 16 | 3414 |
SL-A11 | ARDLRGYC-GLV-CYHDAFDI | GTYDGATQLPV | 19 | 165 |
SL-B10 | ARDLRGYCSWSRLCYHDAFDI | AAYDASRV-SV | 21 | 107 |
SL-D3 | ARDLRGYCAWSRACYHDAFDI | AAWDSSPA-VV | 21 | 412 |
SL-E11 | ARDLRGYC-GVV-CYHDAFDI | GTYDGTERVYV | 19 | 275 |
SL-F5 | ARDLRGYCAWSRSCYHDAFDI | AAWDSRAA-LV | 21 | 495 |
SL-G7 | ARDLRGYC-GVF-CYHDAFDI | GTYDGSASEPV | 19 | 1948 |
2.2. Anti-Fc Antibodies with pH-Dependent Binding Properties
2.2.1. Construction of a Synthetic Library Biased Towards Histidine
2.2.2. Selections against Human Fc Fragments
2.2.3. Characterization of pH-Dependent Anti-hFc Candidates
3. Experimental Section
3.1. Reagents
3.2. Construction of the Synthetic Long Library and Sequencing
3.3. Phage Display Selections against HRP
3.4. Screening ELISA on HRP
3.5. Dose Response Functional Assay of HRP Activity
3.6. Construction of Histidine-Enriched Libraries
3.7. pH-Dependent Phage Display Selections and Screening
3.8. Expression and Purification of Antibodies
3.9. Characterization of pH-Dependent Anti-hFc IgGs
4. Conclusions
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Bonvin, P.; Venet, S.; Kosco-Vilbois, M.; Fischer, N. Purpose-Oriented Antibody Libraries Incorporating Tailored CDR3 Sequences. Antibodies 2015, 4, 103-122. https://doi.org/10.3390/antib4020103
Bonvin P, Venet S, Kosco-Vilbois M, Fischer N. Purpose-Oriented Antibody Libraries Incorporating Tailored CDR3 Sequences. Antibodies. 2015; 4(2):103-122. https://doi.org/10.3390/antib4020103
Chicago/Turabian StyleBonvin, Pauline, Sophie Venet, Marie Kosco-Vilbois, and Nicolas Fischer. 2015. "Purpose-Oriented Antibody Libraries Incorporating Tailored CDR3 Sequences" Antibodies 4, no. 2: 103-122. https://doi.org/10.3390/antib4020103