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Antibodies to MMP-Cleaved Aggrecan

  • Amanda J. Fosang
  • Karena Last
  • David C. Jackson
  • Lorena Brown
Part of the Methods in Molecular Biology™ book series (MIMB, volume 151)

Abstract

The matrix metalloproteinases (MMPs) have a pivotal role in both normal and pathological turnover of the extracellular matrix. Whereas MMP protein can easily be detected by immunolocalization or Western blot analysis, the determination of whether or not an MMP is active and acting on a particular substrate has been more difficult. If the primary sequence and unique cleavage sites within the substrate are known, one means of unambiguously identifying MMP activity is to use antibodies with a unique specificity for antigenic determinants on the newly created N- or C- termini of the degradation products. By definition, these “neoepitope” antibodies recognize a terminal sequence exclusively and do not recognize the same sequence of amino acids located internally as part of the intact protein. This approach has been extremely successful for detecting MMP-derived fragments of aggrecan (the large cartilage proteoglycan) (1, 2, 3, 4, 5) and collagen (6), since aggrecan and collagen are abundant in cartilage matrix, and their neoepitopes are correspondingly abundant. Often MMPs are not the only pro-teinases involved in tissue remodeling and in this situation neoepitope antibodies allow fragments derived from MMPs and other proteinases to be distinguished from each other and compared. In cartilage, aggrecan degradation is mediated by both MMPs, and aggrecanase (7,8) which are members of the ADAMTS family of proteinases (9). The production and use of neoepitope antibodies for investigating cartilage catabolism was first described by Hughes et al. (10). Thereafter neoepitope antibodies were quickly recognized as ideal tools for resolving the products of separate degradative pathways involved in aggrecanolysis.

Keywords

High Performance Liquid Chromatography Ascites Fluid Culture Fluid Booster Injection Fmoc Group 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Humana Press Inc. 2001

Authors and Affiliations

  • Amanda J. Fosang
    • 1
  • Karena Last
    • 1
  • David C. Jackson
    • 2
  • Lorena Brown
    • 2
  1. 1.Department of PaediatricsUniversity of Melbourne, Orthopaedic Research Unit, Royal Children’s HospitalParkvilleAustralia
  2. 2.Department of Microbiology and ImmunologyUniversity of MelbourneParkvilleAustralia

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