Journal: Cartilage

Article Title: A Comparative Study of Fibronectin Cleavage by MMP-1, -3, -13, and -14

doi: 10.1177/1947603511435273

Figure Lengend Snippet: Fibronectin (FN) cleavage by MMP-1, -3, -13, and -14 at neutral pH 7.5. The fibronectin fragments (FN-fs) were measured at time intervals of 0.1, 2, 4, and 24 hours from Western blot analysis of FN digestion by each MMP. The molecular weights were quantified by Totallab TL100 densitometry (Nonlinear Dynamics, Durham, NC) based on intact FN (220 kDa) and FN-fs (70 kDa and 30 kDa) standards of known molecular weight incubated without MMPs for 24 hours (lane 1, CTRL). ( A ) Western blot analysis using a mouse monoclonal antibody specific for an epitope located within the fourth-type 3 repeat of human plasma FN. ( B ) Western blot analysis using a rabbit polyclonal anti-human plasma FN antibody. ( C ) The disappearance of the intact FN subunit expressed as a percentage of the FN remaining. All MMPs cleaved the intact FN by 24 hours.

Article Snippet: Membranes were then washed several times with TBST buffer and treated with either a 10,000-fold dilution of rabbit anti-mouse IgG antibody conjugated to horseradish peroxidase (Santa Cruz Biotechnology, Santa Cruz, CA) in TBST buffer for use with the mouse monoclonal anti-FN antibody or a 7,000-fold dilution of goat anti-rabbit IgG antibody conjugated to horseradish peroxidase (Santa Cruz Biotechnology) in TBST buffer for use with the rabbit polyclonal anti-FN antibody.

Techniques: Western Blot, Molecular Weight, Incubation, Clinical Proteomics

Journal: Cartilage

Article Title: A Comparative Study of Fibronectin Cleavage by MMP-1, -3, -13, and -14

doi: 10.1177/1947603511435273

Figure Lengend Snippet: Fibronectin (FN) cleavage by MMP-1, -3, -13, and -14 at acidic pH 5.5. The fibronectin fragments (FN-fs) were measured at time intervals of 0.1, 2, 4, and 24 hours from Western blot analysis of FN digestion by each MMP. The molecular weights were quantified by Totallab TL100 densitometry (Nonlinear Dynamics, Durham, NC) based on intact FN (220 kDa) and FN-fs (70 kDa and 30 kDa) standards of known molecular weight incubated without MMPs for 24 hours (lane 1, CTRL). ( A ) Western blot analysis using a mouse monoclonal antibody specific for an epitope located within the fourth-type 3 repeat of human plasma FN. ( B ) Western blot analysis using a rabbit polyclonal anti-human plasma FN antibody. ( C ) The disappearance of the intact FN subunit expressed as a percentage of the FN remaining. No FN-fs were detected after FN incubation with MMP-1 and -14.

Article Snippet: Membranes were then washed several times with TBST buffer and treated with either a 10,000-fold dilution of rabbit anti-mouse IgG antibody conjugated to horseradish peroxidase (Santa Cruz Biotechnology, Santa Cruz, CA) in TBST buffer for use with the mouse monoclonal anti-FN antibody or a 7,000-fold dilution of goat anti-rabbit IgG antibody conjugated to horseradish peroxidase (Santa Cruz Biotechnology) in TBST buffer for use with the rabbit polyclonal anti-FN antibody.

Techniques: Western Blot, Molecular Weight, Incubation, Clinical Proteomics