We have located links that may give you full text access.
Comparative Study
Journal Article
Research Support, Non-U.S. Gov't
Multifunctional g3p-peptide tag for current phage display systems.
Journal of Immunological Methods 1998 March 16
We have previously described a monoclonal antibody (mAb), 10C3, directed against the gene-3 protein (g3p) of filamentous phage M13, which was produced to study g3p fusion protein expression in Escherichia coli and its incorporation in the phage capsid [Tesar, M., Beckmann, C., Röttgen, P., Haase, B., Faude, U., Timmis, K., 1995. Monoclonal antibody against pIII of filamentous phage: an immunological tool to study pIII fusion protein expression in phage display systems. Immunology 1, 53-54]. In this study we report mapping of the antigenic epitope of the mAb 10C3, by means of short overlapping peptide-sequences [Frank, R., Overwin, H., 1996. Spot synthesis. In: Morris, G.E. (Ed.), Methods in Molecular Biology, Vol. 66: Epitope Mapping Protocols. Humana Press, Totowa, NJ, pp. 149-169.] comprising the C-terminal half of the g3-protein. A minimal recognizable peptide was found which is represented in the 11 amino acid sequence from positions 292 to 302 of g3p [Wezenbeek van, P.M.G.P., Hulsebos, T.J.M., Schoenmakers, J.G.G., 1980. Nucleotide sequence of the filamentous bacteriophage M13 DNA genome: comparison with phage fd. Gene 11, 129-148]. In order to use the antibody also for detection and purification of recombinant proteins, such as single chain antibodies, the epitope was introduced as a tag sequence into the phagemid pHEN1 [Hoogenboom, H.R., Griffith, A.D., Johnson, K., Chiswell, D.J., Hudson, P., Winter, G., 1991. Multi-subunit proteins on the surface of the filamentous phage: methodologies for displaying antibody (Fab) heavy and light chains. Nucleic Acid Res. 19, 4133-4137; Nissim, A., Hoogenboom, H.R., Tomlinson, I.M., Flynn, G., Midgley, C., Lane, D., Winter, G., 1994. Antibody fragments from a single pot phage display library as immunochemical reagents. EMBO J. 13 (3) 692-698]. Purified single chain antibodies containing this tag were detectable down to a concentration of 2 ng ml(-1) under non-denaturing conditions (ELISA) or 4 ng per lane on immunoblots. The high sensitivity of the antibody for the peptide tag was reflected in the antibody affinity constant K(D) of 6.80 x 10(-10) M, which was determined by real time biomolecular interaction analysis (BIA) based on surface plasmon resonance (SPR) [Karlsson, R., Fält, A., 1997. Experimental design for kinetic analysis of protein-protein interactions with surface plasmon resonance biosensors. J. Immunol. Methods 200, 121-133]. Finally, recombinant proteins in E. coli periplasmic extracts could be purified in a single step by affinity purification using immobilized mAb 10C3. These studies demonstrated that the new peptide-tag and its corresponding mAb represents a versatile tool for the detection of recombinant proteins selected by phage display technology.
Full text links
Related Resources
Trending Papers
Challenges in Septic Shock: From New Hemodynamics to Blood Purification Therapies.Journal of Personalized Medicine 2024 Februrary 4
Molecular Targets of Novel Therapeutics for Diabetic Kidney Disease: A New Era of Nephroprotection.International Journal of Molecular Sciences 2024 April 4
The 'Ten Commandments' for the 2023 European Society of Cardiology guidelines for the management of endocarditis.European Heart Journal 2024 April 18
A Guide to the Use of Vasopressors and Inotropes for Patients in Shock.Journal of Intensive Care Medicine 2024 April 14
Get seemless 1-tap access through your institution/university
For the best experience, use the Read mobile app
All material on this website is protected by copyright, Copyright © 1994-2024 by WebMD LLC.
This website also contains material copyrighted by 3rd parties.
By using this service, you agree to our terms of use and privacy policy.
Your Privacy Choices
You can now claim free CME credits for this literature searchClaim now
Get seemless 1-tap access through your institution/university
For the best experience, use the Read mobile app