Abstract
With the increasing need for understanding antibody specificity in antibody and vaccine research, pepscan assays provide a rapid method for mapping and profiling antibody responses to continuous epitopes. We have developed a relatively low-cost method to generate peptide microarray slides for studying antibody binding. Using a setup of an IntavisAG MultiPep RS peptide synthesizer, a Digilab MicroGrid II 600 microarray printer robot, and an InnoScan 1100 AL scanner, the method allows the interrogation of up to 1536 overlapping, alanine-scanning, and mutant peptides derived from the target antigens. Each peptide is tagged with a polyethylene glycol aminooxy terminus to improve peptide solubility, orientation, and conjugation efficiency to the slide surface.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Plotkin SA (2010) Correlates of protection induced by vaccination. Clin Vaccine Immunol 17:1055–1065
Strohl WR (2009) Therapeutic monoclonal antibodies: past, present and future. In: An Z (ed) Therapeutic monoclonal antibodies: from bench to clinic. Wiley, Hoboken, NJ, pp 3–50
Garces F, Sok D, Kong L, McBride R, Kim HJ, Saye-Francisco KF, Julien JP, Hua Y, Cupo A, Moore JP, Paulson JC, Ward AB, Burton DR, Wilson IA (2014) Structural evolution of glycan recognition by a family of potent HIV antibodies. Cell 159:69–79
Moore JP, Sodroski J (1996) Antibody cross-competition analysis of the human immunodeficiency virus type 1 gp120 exterior envelope glycoprotein. J Virol 70:1863–1872
Law M, Maruyama T, Lewis J, Giang E, Tarr AW, Stamataki Z, Gastaminza P, Chisari FV, Jones IM, Fox RI, Ball JK, McKeating JA, Kneteman NM, Burton DR (2008) Broadly neutralizing antibodies protect against hepatitis C virus quasispecies challenge. Nat Med 14:25–27
Giang E, Dorner M, Prentoe JC, Dreux M, Evans MJ, Bukh J, Rice CM, Ploss A, Burton DR, Law M (2012) Human broadly neutralizing antibodies to the envelope glycoprotein complex of hepatitis C virus. Proc Natl Acad Sci U S A 109:6205–6210
Kong L, Giang E, Nieusma T, Kadam RU, Cogburn KE, Hua Y, Dai X, Stanfield RL, Burton DR, Ward AB, Wilson IA, Law M (2013) Hepatitis C virus E2 envelope glycoprotein core structure. Science 342:1090–1094
Janvier B, Archinard P, Mandrand B, Goudeau A, Barin F (1990) Linear B-cell epitopes of the major core protein of human immunodeficiency virus types 1 and 2. J Virol 64:4258–4263
Tomaras GD, Binley JM, Gray ES, Crooks ET, Osawa K, Moore PL, Tumba N, Tong T, Shen X, Yates NL, Decker J, Wibmer CK, Gao F, Alam SM, Easterbrook P, Abdool Karim S, Kamanga G, Crump JA, Cohen M, Shaw GM, Mascola JR, Haynes BF, Montefiori DC, Morris L (2011) Polyclonal B cell responses to conserved neutralization epitopes in a subset of HIV-1-infected individuals. J Virol 85:11502–11519
Li XQ, Qiu LW, Chen Y, Wen K, Cai JP, Chen J, Pan YX, Li J, Hu DM, Huang YF, Liu LD, Ding XX, Guo YH, Che XY (2013) Dengue virus envelope domain III immunization elicits predominantly cross-reactive, poorly neutralizing antibodies localized to the AB loop: implications for dengue vaccine design. J Gen Virol 94:2191–2201
Zhao S, Qi T, Guo W, Lu G, Xiang W (2013) Identification of a conserved B-cell epitope in the equine arteritis virus (EAV) N protein using the pepscan technique. Virus Genes 47:292–297
Ruwona TB, Giang E, Nieusma T, Law M (2014) Fine mapping of murine antibody responses to immunization with a novel soluble form of hepatitis C virus envelope glycoprotein complex. J Virol 88:10459–10471
Uchtenhagen H, Schiffner T, Bowles E, Heyndrickx L, LaBranche C, Applequist SE, Jansson M, De Silva T, Back JW, Achour A, Scarlatti G, Fomsgaard A, Montefiori D, Stewart-Jones G, Spetz AL (2014) Boosting of HIV-1 neutralizing antibody responses by a distally related retroviral envelope protein. J Immunol 192:5802–5812
Ruwona TB, McBride R, Chappel R, Head SR, Ordoukhanian P, Burton DR, Law M (2014) Optimization of peptide arrays for studying antibodies to hepatitis C virus continuous epitopes. J Immunol Methods 402:35–42
IntavisAG—MultiPep RS: Parallel Peptide Synthesis, User Manual
Winkler DF, Hilpert K, Brandt O, Hancock RE (2009) Synthesis of peptide arrays using SPOT-technology and the CelluSpots-method. Methods Mol Biol 570:157–174
Hilpert K, Winkler DF, Hancock RE (2007) Peptide arrays on cellulose support: SPOT synthesis, a time and cost efficient method for synthesis of large numbers of peptides in a parallel and addressable fashion. Nat Protoc 2:1333–1349
Busch J, McBride R, Head SR (2010) Production and application of glycan microarrays. Methods Mol Biol 632:269–282
Acknowledgement
This work was supported by NIH grants AI079031 and AI106005 (M.L.). This is TSRI manuscript number 28094.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2016 Springer Science+Business Media New York
About this protocol
Cite this protocol
McBride, R., Head, S.R., Ordoukhanian, P., Law, M. (2016). Low-Cost Peptide Microarrays for Mapping Continuous Antibody Epitopes. In: Cretich, M., Chiari, M. (eds) Peptide Microarrays. Methods in Molecular Biology, vol 1352. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3037-1_6
Download citation
DOI: https://doi.org/10.1007/978-1-4939-3037-1_6
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-3036-4
Online ISBN: 978-1-4939-3037-1
eBook Packages: Springer Protocols