The "reverse capture" autoantibody microarray: An innovative approach to profiling the autoantibody response to tissue-derived native antigens

Ehrlich, Joshua R., Tang, Liangdan, Caiazzo Jr., Robert J., Cramer, Daniel W., Ng, Shu-Kay, Ng, Shu-Wing and Liu, Brian C.-S. (2008) The "reverse capture" autoantibody microarray: An innovative approach to profiling the autoantibody response to tissue-derived native antigens. Methods in Molecular Biology, 441 175-192. doi:10.1007/978-1-60327-047-2_12


Author Ehrlich, Joshua R.
Tang, Liangdan
Caiazzo Jr., Robert J.
Cramer, Daniel W.
Ng, Shu-Kay
Ng, Shu-Wing
Liu, Brian C.-S.
Title The "reverse capture" autoantibody microarray: An innovative approach to profiling the autoantibody response to tissue-derived native antigens
Journal name Methods in Molecular Biology   Check publisher's open access policy
ISSN 1064-3745
1940-6029
ISBN 9781588296795
Publication date 2008
Sub-type Article (original research)
DOI 10.1007/978-1-60327-047-2_12
Open Access Status
Volume 441
Start page 175
End page 192
Total pages 18
Place of publication New York, NY United States
Publisher Humana Press
Subject 1312 Molecular Biology
1311 Genetics
Abstract Recently, we reported the development and use of a "reverse capture" antibody microarray for the purpose of investigating antigen-autoantibody profiling. This platform was developed to allow researchers to characterize and compare the autoantibody profiles of normal and diseased patients. Our "reverse capture" protocol is based on the dual-antibody sandwich immunoassay of enzyme-linked immunosorbent assay (ELISA), and we have previously reported its use to detect autoimmunity to epitopes found on native antigens derived from tumor cell lines. In this protocol, we used ovarian cancer as a model system to adapt the "reverse capture" procedure for use with native antigens derived from frozen tissue samples. The use of this platform in studies of autoimmunity is valuable because it allows for the detection of autoantibody reactivity with epitopes found on the post-translational modifications (PTMs) of native antigens, a feature not present with other protein array platforms. In the first step in the "reverse capture" process, tissue-derived native antigens are immobilized onto the 500 monoclonal antibodies that are spotted in duplicate on the array surface. Using the captured antigens as "baits," we then incubate the array with labeled IgG from test and control samples, and perform a two-slide dye-swap to account for any dye effects. Here, we present a detailed description of the "reverse capture" autoantibody microarray for use with tissue-derived native antigens.
Keyword Autoantibodies
Autoantigens
Ovarian cancer
Protein microarrays
Tissues
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status UQ

Document type: Journal Article
Sub-type: Article (original research)
Collection: School of Mathematics and Physics
 
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