SpringerProtocols: Abstract: Methods for Measuring Type I Collagen Synthesis In Vitro

Methods for Measuring Type I Collagen Synthesis In Vitro

By: David C. Rishikof⁴, Ping-Ping Kuang⁴, Mangalalaxmy Subramanian⁴, Ronald H. Goldstein⁴

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The excess accumulation of type I collagen within tissues leads to organ dysfunction and occurs as a result of an imbalance between synthesis and degradation. This chapter outlines several methods to assess the in vitro production of type I collagen that are employed in our laboratory. We describe Western immunoblotting of intact α1(I) collagen using antibodies directed to α1(I) collagen amino and carboxyl propeptides. The measurement of α1(I) collagen mRNA levels using real-time polymerase chain reaction is then outlined. Finally, methods to determine the transcriptional regulation of α1(I) collagen using a nuclear run-on assay are described.

Affiliation(s): (4) The Pulmonary Center, Boston University School of Medicine, The Boston VA Medical Center, Boston, MA

Book Title: Fibrosis Research: Methods and Protocols

Series: Methods in Molecular Medicine | Volume: 117 | Pub. Date: Aug-08-2005 | Page Range: 129-140 | DOI: 10.1385/1-59259-940-0:129

Subject: Molecular Medicine

Key Words: Type I collagen - Western immunoblot - real-time PCR - nuclear run-on assay

References

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