-Development of the first phage display systems for the display of large proteins, particularly 50 KD heterodimeric antibody Fab fragments
-Development of the first phage displayed Human Antibody Libraries
-Cloning of the first High Affinity Human Monoclonal Antibodies using Phage Display
-Development of Methodologies for the construction and selection of phage display libraries of Human, Mouse, Rabbit and Chicken antibody Fab’s and scFv’s
-Development of the first Synthetic Antibody Libraries
-Development of the first Zinc Finger Display Libraries
-Development of the first Motif Grafted Antibodies
-Development of one the most potent HIV-1 neutralizing antibodies known, IgG-b12
-Development of robust methods to improve Antibody affinity- CDR Walking
-Development of the first in vitro evolved Antibodies with enhanced biological activity, the anti-HIV-1 antibody 3B3
-Development of phage-based methods for the humanization of mouse and rabbit Antibodies
-Development of a wide range of anti-Cancer Antibodies
-Development of novel Catalytic Antibodies and Catalytic peptides

Barbas, C. F., III; Burton, D. R.; Scott, J.K., Silverman, G.J. Eds. (2001) Phage Display: A Laboratory Manual; Cold Spring Harbor Laboratory Press: Cold Spring Harbor, New York, 736 pages. Information about the manual can be found by clicking on the title or by purchasing directly from www.cshlpress.com or www.Amazon.com.

 

1. pComb3X Set – newest version of pComb3 (pComb3XSS, pComb3XTT, pComb3XLambda)
2. pComb3H Set – 2nd generation of pComb3 (pComb3HSS, pComb3HTT, pAraH6HATT)
3. The original pCombs (pComb3, pComb8)
4. Miscellaneous (pJF3H, pIgG)

 

pComb3X Family (Genbank AF268281) Maps

 

pComb3H Family (Genebank AF268280) Maps
pComb3HSS Text
pComb3HTT Text

pAraH6HATT Text
pMalCSS (cytoplasmic expression) Text
pMalPSS (periplasmic expression) Text

The Original pComb

pComb3 Text & Map
pComb8 Text & Map

The original pComb3 vector was designed for phage display of Fabs which are cloned in one chain at a time using SacI/XbaI restriction sites (for the light chain) and XhoI/SpeI restriction sites (for the heavy chain). This vector system has also been used for the display of a wide variety of other proteins like zinc fingers, peptides, and cDNA fragments. Phage can be produced which express Fab or other proteins or peptides of interest fused to the phage pIII protein for expression on the head of the phage. Soluble Fab can be expressed by removing the gene for the pIII phage fusion protein by SpeI/NheI digest. Sufficient soluble protein is also found in the periplasmic space as a result of proteolysis. pComb8 is nearly identical but contains the phage pVIII fusion protein for multi-valent expression along the sides of the phage via fusion with pVIII.

Select References:

Barbas III, C.F. (1995) Synthetic Human Antibodies. Nature Medicine 1,837-839.

Barbas III, C.F., Bjorling, E., Chiodi, F., Dunlop, N., Cababa, D., Jones, T.M., Zebedee, S.L., Persson, M.A.A., Nara, P.L., Norrby, E. and Burton, D.R. (1992) Recombinant Human Fab fragments neutralize human type 1 immunodeficiency virus in vitro. Proc. Natl. Acad. Sci. USA 89, 9339-9343

Barbas III, C.F., Kang, A.S., Lerner, R.A. and Benkovic, S.J. (1991) Assembly of combinatorial antibody libraries on phage surfaces: The gene III site. Proc. Natl. Acad. Sci. USA 88, 7978-7982

Barbas III, C.F., Crowe, J.E., Cababa, D., Jones, T.M., Zebedee, S.L., Murphy, B.R., Chanock, R.M. and Burton, D.R. (1992) Human Monoclonal Fab Fragments derived from a combinatorial library bind to respiratory syncytial virus F glycoprotein and neutralize infectivity. Proc. Natl. Acad. Sci. USA 89, 10164-10168

Barbas III, C.F., Persson, M.A.A., Koenig, S., Chanock, R.M., Burton, D.R. and Lerner, R.A. (1992) A large array of human monoclonal antibodies to HIV-1 from combinatorial libraries of an asymptomatic seropositive individual. Vaccines '92: Modern Approaches to New Vaccines including Prevention of AIDS, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York , 9-12

Barbas III, C.F., Bain, J.D., Hoekstra, D.M. and Lerner, R.A. (1992) Semi-synthetic combinatorial antibody libraries: A chemical solution to the diversity problem. Proc. Natl. Acad. Sci. USA, 89:4457-4461.

Lerner, R.A., Kang, A.S., Bain, J.D., Burton, D.R., Barbas III, C.F. (1992) Antibodies without immunization. Science 258:1313-1314.

Barbas III, C.F., Amberg, W., Somincsits, A., Jones, T.M. and Lerner, R.A. (1993) Selection of human anti-hapten antibodies from semisynthetic libraries. Gene, 137:57-62.

Burton, D.R., Barbas III, C.F., Persson, M.A.A., Koenig, S., Chanock, R.M. and Lerner, R.A. (1991) A large array of human monoclonal antibodies to type 1 human immunodeficiency virus from combinatorial libraries of asymptomatic seropositive individuals. Proc. Natl. Acad. Sci. USA 88, 10134-10137

Barbas III, C.F., Languino, L.R. and Smith, J.W. (1993) High Affinity Self-Reactive Human Antibodies by Design and Selection: Targeting the Integrin Ligand Binding Site, Proc. Natl. Acad. Sci. USA 90:10003-10007.

Smith, J.W., Hu, D., Satterthwait, A.C., Pinz-Sweeney, S. and Barbas III, C.F. (1994) Building Synthetic Antibodies as Adhesive Ligands for Integrins. J. Biol. Chem. 269:32788-32795.

Barbas III, C.F., and Wagner, J. (1995) Synthetic Human Antibodies: Selecting and Evolving Functional Proteins. Methods, A Companion to Methods in Enzymology 8, 94-103.

 

Other Related Vectors

pJF3H Text & Map

Modified version of pComb3H containing Jun, Fos and a cloning site for cDNA expression. Developed in the lab of Gregg Silverman at UCSD.

References:

Barbas, C. F., III; Burton, D. R.; Scott, J.K., Silverman, G.J. Eds. (2001) Phage Display: A Laboratory Manual;
Cold Spring Harbor Laboratory Press: Cold Spring Harbor, New York, , 736 pages.

pIgG Text & Map

Vector for conversion of Fab to IgG for expression in mammalian cells.

References:

Rader, C; Popkov, M.; Neves, J.A.; Barbas III, C.F. (2002) Integrin avb3 Targeted Therapy of Kaposi’s Sarcoma with an In Vitro Evolved Antibody . FASEB, 16(14), 2000-2002.

 

Phage Display: A Laboratory Manual
By Carlos F. Barbas III, The Scripps Research Institute;
Dennis R. Burton, The Scripps Research Institute;
Jamie K. Scott, Simon Fraser University;
Gregg J. Silverman, University of California, San Diego
2001, 736 pp., illus., index
Paperback $75 ISBN 0-87969-546-3

*Customers in Europe, the Middle East, and Africa should check for this publication at the CSHL Press Europe website.

Chapter 1. Filamentous Phage Biology
Chapter 2. Phage-display Vectors
Chapter 3. Antibody Libraries
Chapter 4. Peptide Libraries
Chapter 5. Functional Domains and Scaffolds
Chapter 6. Gene Fragment Libraries and Genomic and cDNA Expression Cloning

Chapter 7. Overview: Amplification of Antibody Genes
Chapter 8. Generation of Antibody Libraries: Immunization, RNA Preparation, and cDNA Synthesis
Chapter 9. Generation of Antibody Libraries: PCR Amplification and Assembly of Light- And
Heavy-chain Coding Sequences
Chapter 10. Selection from Antibody Libraries
Chapter 11. Analysis of Selected Antibodies
Chapter 12. Production and Purification of Fab and scFv
Chapter 13. Antibody Engineering

Chapter 14. Overview: Peptide Libraries
Chapter 15. General Phage Methods
Chapter 16. Production of Peptide Libraries
Chapter 17. Screening Peptide Libraries
Chapter 18. Analysis of Phage-borne Peptides
Chapter 19. Construction and Use of pIII-displayed Peptide Libraries

Chapter 20. Construction and Selection from Gene Fragment Phage-display Expression Libraries
Chapter 21. Construction and Selection from cDNA Phage-display Expression Libraries
Chapter 22. In Vivo Selection of Phage-display Libraries
Chapter 23. Cell-surface Selection and Analysis of Monoclonal Antibodies from Phage Libraries

Appendix 1. Useful Information
Appendix 2. Recipes
Appendix 3. General Procedures
Appendix 4. Cautions
Appendix 5. Suppliers
Appendix 6. Trademarks