Search:        >> Advanced Search
Guidelines | Subscriptions | About | exPRESS - Current - Archive | Business Information | Contact

This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Brown, J. K. Articles by Miller, H. R.P. Search for Related Content PubMed PubMed Citation Articles by Brown, J. K. Articles by Miller, H. R.P. Social Bookmarking                      What's this?

Primary Antibody–Fab Fragment Complexes : A Flexible Alternative to Traditional Direct and Indirect Immunolabeling Techniques

Jeremy K. Brown, Alan D. Pemberton, Steven H. Wright and Hugh R.P. Miller

Department of Veterinary Clinical Studies, University of Edinburgh, Easter Bush Veterinary Centre, Midlothian, United Kingdom

Correspondence to: Jeremy K. Brown, Dept. of Veterinary Clinical Studies, University of Edinburgh, Easter Bush Veterinary Centre, Midlothian EH25 9RG, UK. E-mail: Jeremy.brown2{at}ed.ac.uk

Immunolabeling with immune complexes of primary and secondary antibodies offers an attractive method for detecting and quantifying specific antigen. Primary antibodies maintain their affinity for specific antigen after labeling with Fab fragments in vitro. Incubation of these immune complexes with excess normal serum from the same species as the primary antibody prevents free Fab fragments from recognizing immunoglobulin. Effectively a hybrid between traditional direct and indirect immunolabeling techniques, this simple technique allows primary antibodies to be non-covalently labeled with a variety of reporter molecules as and when required. Using complexes containing Fab fragments that recognize both the Fc and F(ab')₂ regions of IgG, we show that this approach prevents nonspecific labeling of endogenous immunoglobulin, can be used to simultaneously detect multiple antigens with primary antibodies derived from the same species, and allows the same polyclonal antibody to be used for both antigen capture and detection in ELISA. (J Histochem Cytochem 52:1219–1230, 2004)

Key Words: endogenous immunoglobulin • Fab • fluorescence • multicolor • multilabeling • flow cytometry • ELISA

CiteULike    Complore    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				R. R. Mundegar, E. Franke, R. Schafer, M. Zweyer, and A. Wernig Reduction of High Background Staining by Heating Unfixed Mouse Skeletal Muscle Tissue Sections Allows for Detection of Thermostable Antigens With Murine Monoclonal Antibodies J. Histochem. Cytochem., November 1, 2008; 56(11): 969 - 975. [Abstract] [Full Text] [PDF]

				J. M. Schmitz, V. J. McCracken, R. A. Dimmitt, and R. G. Lorenz Expression of CXCL15 (Lungkine) in Murine Gastrointestinal, Urogenital, and Endocrine Organs J. Histochem. Cytochem., May 1, 2007; 55(5): 515 - 524. [Abstract] [Full Text] [PDF]

				D. von Smolinski, M. Blessenohl, C. Neubauer, K. Kalies, and A. Gebert Validation of a Novel Ultra-Short Immunolabeling Method for High-Quality mRNA Preservation in Laser Microdissection and Real-Time Reverse Transcriptase-Polymerase Chain Reaction J. Mol. Diagn., May 1, 2006; 8(2): 246 - 253. [Abstract] [Full Text] [PDF]

Guidelines | Subscriptions | About | exPRESS - Current - Archive | Business Information | Contact

The Journal of Histochemistry & Cytochemistry is owned, published, and licensed by The Histochemical Society © 2004