Rat TIMP-1 Antibody

Detection of Rat TIMP-1 by Western Blot

Preconditioning with HGF induces Timp1 (tissue inhibitor of metalloproteinases-1) protein synthesis, which is followed by Stat3 phosphorylation and protection against Doxo-induced apoptosis. (a) H9c2 cardiomyoblasts were treated with Doxo (25 μM) or Doxo+HGF (0.5 nM). For cell treatments, see Figure 1a. Timp1 protein levels were evaluated by Western blotting at different points of recovery time (a). Cells were also treated with (b) 500 nM JNJ38877605 Met inhibitor (Doxo+HGF+JNJ) or (c,d) 1 µM PD98059 Erk1,2 inhibitor (Doxo+HGF+PD). Protein (b,d) and mRNA (c) levels of Timp1 were analyzed after 24 h of recovery time. Polr2a was used as reference gene for the expression data normalization. *** p < 0.005 significantly different from Doxo-treated cells. (e,f) H9c2 cells were pretreated with HGF alone, or with HGF+Cycloheximide (CHX, 10 μM) for 4 h. Then, Doxo (25 μM) was added in the last 1 h. The CHX treatment was performed 30 min before adding HGF and was maintained during all the treatment protocol. Protein levels of Timp1, P-Stat3 (Y705), Stat3, P-Erk1,2, Erk2 (e) and total and cleaved caspase 3 (f) were detected at different time points of recovery time (0, 2 and 6 h). (g,h) H9c2 cells were treated with Doxo (25 μM), Doxo+HGF (0.5 nM) or Doxo+HGF+Timp1 siRNA. Protein levels of Timp1, P-Stat3 (Y705), Stat3, P-Erk1,2 and Erk2 (g) and gammaH2AX, H2AX and cleaved/total caspase 3 ratios (h) were measured after 24 h of recovery time. The ratios calculated between cleaved and total caspase 3 are shown. alphatubulin was used as the loading control in all Western blots. Data are representative results of three independent experimental replicates. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/32722178), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of TIMP-1 in Rat Brain.

TIMP-1 was detected in perfusion fixed paraffin-embedded sections of Rat Brain using Goat Anti-Rat TIMP-1 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF580) at 5 µg/mL for 1 hour at room temperature followed by incubation with the Anti-Goat IgG VisUCyte™ HRP Polymer Antibody (Catalog # VC004). Before incubation with the primary antibody, tissue was subjected to heat-induced epitope retrieval using VisUCyte Antigen Retrieval Reagent-Basic (Catalog # VCTS021). Tissue was stained using DAB (brown) and counterstained with hematoxylin (blue). Specific staining was localized to neuronal cell bodies. View our protocol for IHC Staining with VisUCyte HRP Polymer Detection Reagents.

Detection of Rat TIMP-1 by Western Blot

Preconditioning with HGF induces Timp1 (tissue inhibitor of metalloproteinases-1) protein synthesis, which is followed by Stat3 phosphorylation and protection against Doxo-induced apoptosis. (a) H9c2 cardiomyoblasts were treated with Doxo (25 μM) or Doxo+HGF (0.5 nM). For cell treatments, see Figure 1a. Timp1 protein levels were evaluated by Western blotting at different points of recovery time (a). Cells were also treated with (b) 500 nM JNJ38877605 Met inhibitor (Doxo+HGF+JNJ) or (c,d) 1 µM PD98059 Erk1,2 inhibitor (Doxo+HGF+PD). Protein (b,d) and mRNA (c) levels of Timp1 were analyzed after 24 h of recovery time. Polr2a was used as reference gene for the expression data normalization. *** p < 0.005 significantly different from Doxo-treated cells. (e,f) H9c2 cells were pretreated with HGF alone, or with HGF+Cycloheximide (CHX, 10 μM) for 4 h. Then, Doxo (25 μM) was added in the last 1 h. The CHX treatment was performed 30 min before adding HGF and was maintained during all the treatment protocol. Protein levels of Timp1, P-Stat3 (Y705), Stat3, P-Erk1,2, Erk2 (e) and total and cleaved caspase 3 (f) were detected at different time points of recovery time (0, 2 and 6 h). (g,h) H9c2 cells were treated with Doxo (25 μM), Doxo+HGF (0.5 nM) or Doxo+HGF+Timp1 siRNA. Protein levels of Timp1, P-Stat3 (Y705), Stat3, P-Erk1,2 and Erk2 (g) and gammaH2AX, H2AX and cleaved/total caspase 3 ratios (h) were measured after 24 h of recovery time. The ratios calculated between cleaved and total caspase 3 are shown. alphatubulin was used as the loading control in all Western blots. Data are representative results of three independent experimental replicates. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/32722178), licensed under a CC-BY license. Not internally tested by R&D Systems.