AffiASSAY® Gelatinase (Gelatin Degradation/Zymography) Assay Kit (Fluorometric)

Specifications

Description

Gelatinases are a type of matrix zinc-dependent metalloproteases (MMPs) that degrade gelatins and a variety of other extracellular matrix proteins. These enzymes are synthesized as latent zymogens that are activated by proteolysis and inhibited by tissue inhibitors of metalloproteases (TIMPs). Two mammalian gelatinases, Gelatinase A (MMP-2) and Gelatinase B (MMP-9), are critical for basement membrane degradation and are highly upregulated in variety of tumor cells. Gelatinase activity is usually detected by small peptide-based activity assays which may suffer from lack of substrate specificity. Other methods for gelatinase activity include gelatin Zymography where samples are electrophoresed on a gelatin-containing SDS-PAGE, and further renatured in a suitable buffer for 12-16 h. The zymogram is subsequently stained, and areas of digestion appear as clear bands against a darkly stained background where the substrate has been degraded by the enzyme. Such methods are laborious, time-consuming and may lead to the loss of enzymatic activity as renaturation may not be completely reversible. BioVision’s Gelatinase Activity Assay Kit utilizes a hybrid approach for the detection of gelatinase activity by employing a highly quenched gelatin substrate which upon cleavage by a suitable gelatinase releases a fluorophore, which can be easily quantified using a conventional microplate reader. This method is substrate-specific, simple, fast, high-throughput adaptable and amenable to the sensitive detection of gelatinase activity (as low as 0.06 mCDU for bacterial collagenase) in biological samples.