Anti-Collagen I + Collagen III 抗体

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen I + Collagen III antibody (AB34710)

Immunohistochemical analysis of formalin-fixed paraffin-embedded human tubuli and blood vessels labelling Collagen I and Collagen III with ab34710 at 1/15 for 1 hour at 37 °C followed by a ready to Polymer-HRP, Rabbit/Mouse Detection Kit. Blocking : Peroxidase-Blocking Solution for 10 minutes. Substrate : DAB-Chromogen, Rabbit/Mouse. Staining/Results : Intense collagen I staining of fibres surrounding tubuli and around blood vessels. Counterstained with  hematoxylin for 15 seconds. Heat induced epitope retrieval (HIER) using Tris-EDTA-citrate buffer pH 7.8 for 5 minutes.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen I + Collagen III antibody (AB34710)

Immunohistochemical analysis of formalin-fixed paraffin-embedded human stomach mucosa (TMA) tissue labelling Collagen I and Collagen III with ab34710 at 1/15 dilution for 1 hour at 37 °C followed by a ready to use Polymer-HRP, Rabbit/Mouse Detection Kit. Blocking : Peroxidase-Blocking Solution for 10 minutes. Substrate : DAB-Chromogen, Rabbit/Mouse. Staining/Results : basement membranes and blood vessels. Counterstained with hematoxylin for 15 seconds. Heat induced epitope retrieval (HIER) using Tris-EDTA-citrate buffer pH 7.8 for 5 minutes.

• IHC-P

Supplier Data

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Collagen I + Collagen III antibody (AB34710)

Immunohistochemical analysis of formalin-fixed paraffin-embedded smooth muscle cells of the human stomach wall labelling Collagen I and Collagen III with ab34710 at 1/15 dilution for 1 hour at 37 °C followed by a ready to use Polymer-HRP, Rabbit/Mouse Detection Kit. Blocking : Peroxidase-Blocking Solution for 10 minutes. Substrate : DAB-Chromogen, Rabbit/Mouse. Staining/Results : smooth muscle cells surrounded by collagen fibers. Counterstained with hematoxylin for 15 seconds. Heat induced epitope retrieval (HIER) using Tris-EDTA-citrate buffer pH 7.8 for 5 minutes.

• WB

Supplier Data

Western blot - Anti-Collagen I + Collagen III antibody (AB34710)

DyLight™ 649 anti-rabbit secondary antibody at 1 : 20,000 for 30 min at RT.

Blocking Buffer for 30 min at room temperature - proprietary protein formulation in TRIS buffered saline at pH 7.6 with thimerosal added as an antimicrobial agent.

Other Band(s) : Collagen Type I splice variants and isoforms.

All lanes:

Western blot - Anti-Collagen I + Collagen III antibody (ab34710) at 1/1000 dilution

All lanes:

Western blot - Native Human Collagen I protein (<a href='/products/proteins-peptides/native-human-collagen-i-protein-ab7533'>ab7533</a>) at 0.05 µg

Secondary

All lanes:

DyLight™ 649 anti-rabbit secondary antibody at 1/20000 dilution

Predicted band size: 139 kDa

false

• WB

Supplier Data

Western blot - Anti-Collagen I + Collagen III antibody (AB34710)

All lanes:

Western blot - Anti-Collagen I + Collagen III antibody (ab34710)

Lane 1:

Human Collagen I Reduced/Boiled at 10 µg

Lane 2:

Human Collagen I Reduced/Non-Boiled at 10 µg

Lane 3:

Human Collagen III Reduced/Boiled at 10 µg

Lane 4:

Human Collagen III Reduced/ Non-Boiled at 10 µg

Lane 6:

Human Collagen I Non-Reduced/Boiled at 10 µg

Lane 7:

Human Collagen I Non-Reduced/Non-Boiled at 10 µg

Lane 8:

Human Collagen III Non-Reduced/Boiled at 10 µg

Lane 9:

Human Collagen III Non-Reduced/ Non-Boiled at 10 µg

Predicted band size: 139 kDa

false

Exposure time: 8.9s

• WB

CiteAb

Western blot - Anti-Collagen I + Collagen III antibody (AB34710)

Western Blotting using Anti-Collagen I antibody, ab34710. Publication image from Jeng, Y. M. et al., 2022, Nat Commun, 35618735. Legend direct from paper.

Direct tumor-fibroblast contact triggers activin A secretion leading to EMT of tumor cells and myofibroblast activation.a Human cytokine array used conditional medium from transwell (indirect, ID) versus direct (D) co-culture of BxPC-3 with pancreatic stellate cells (PSCs). Cytokine levels in direct co-cultured condition were normalized to indirect co-cultured condition. b Quantitative real-time PCR analyses of CLDN1, VIM, Snail, Twist1, ZEB1, and ZEB2 expression were performed in BxPC-3 cells treated with recombinant human Activin A (50 and 100 ng/ml) compared to untreated cells. Values were presented as mean ± SD (n = 3). c Western blotting analysis of proteins harvested from BxPC-3 cells treated with recombinant human activin A (50 and 100 ng/ml) compared to untreated cells. d Sphere-formation assay of BxPC-3 cells treated with recombinant human activin A compared to untreated cells. Values were presented as mean ± SD (n = 4). e Western blotting analysis of proteins harvested from mono-cultured and direct co-cultured (D) tumor cells (BxPC-3-RFP) treated with IgG (1 µg/ml) or anti-activin A-neutralizing antibody (1 µg/ml, 693604, Biolegend). FACS was performed to collect RFP+ tumor cells after 48 h of mono- or co-culture conditions. f Sphere-formation assay of BxPC-3-RFP cells in mono-cultured and direct co-cultured conditions treated with IgG (2 µg/ml) or anti-activin A-neutralizing antibody (1 and 2 µg/ml). Values were presented as mean ± SD (n = 4). g, h 3D spheroid invasion assay of BxPC-3-RFP spheroids in mono-cultured and direct co-cultured conditions treated with IgG (4 µg/ml) or anti-activin A-neutralizing antibody (2 and 4 µg/ml). g Representative images. Scale bar, 200 µm. h Extensive rate of each tumor spheroid was calculated by normalizing RFP+ area at 72 h to RFP+ area at 1 h (left panel) and by normalizing RFP+ maximal diameter at 72 h to RFP+ maximal diameter at 1 h (right panel). Values were presented as mean ± SD (n = 9). i, j Subcutaneous injection of BxPC-3 cells and mixed fibroblasts/BxPC-3 cells treated with IgG (1 µg/ml) or anti-activin A-neutralizing antibody (1 µg/ml) every 3 days and collected tumor samples at day 14 after inoculation. Representative images i and quantification results of tumor weight j. Scale bar, 1 cm. Values were presented as mean ± SD (n = 6). k, l Subcutaneous injection of BxPC-3 cells and mixed fibroblasts/BxPC-3 cells treated with recombinant human follistatin 20 ng/ml every 3 days and collected tumor samples at day 14 after inoculation. Representative images k and quantification results of tumor weight l. Scale bar, 1 cm. Values were presented as mean ± SD (n = 5). m, n Quantitative real-time PCR analyses and western blotting analysis of ACTA2 and COL1A1 expression were performed in PSCs treated with recombinant human activin A (50 and 100 ng/ml) compared to untreated cells for 24 h and 48 h, respectively. For qPCR analyses, values were presented as mean ± SD (n = 3). o, p Quantitative real-time PCR analyses and western blotting analysis of ACTA2 and COL1A1 expression in mono-cultured, indirect co-cultured (ID), and direct co-cultured (D) fibroblasts (PSC-GFP) with tumor cells (BxPC-3). FACS was performed to collect GFP+ fibroblasts after 24 h and 48 h for mRNA and protein samples, respectively. For qPCR analyses, values were presented as mean ± SD (n = 3). q Western blotting analysis of proteins harvested from mono-cultured and direct co-cultured (D) fibroblasts (PSC-GFP) treated with IgG (2 µg/ml) or anti-activin A-neutralizing antibody (1 and 2 µg/ml). FACS was performed to collect GFP+ fibroblasts after 48 h of mono- or co-cultured conditions. r, s BxPC-3 cells and mixed PSCs/BxPC-3 cells treated with IgG (2 µg/ml) or anti-activin A-neutralizing antibody (2 µg/ml) every 3 days and collected tumor samples at day 14 after inoculation. Representative IHC images r and H Score s ofαSMA expression. Scale bar, 0.5 mm. Values were presented as mean ± SD (n = 6). t, u BxPC-3 cells and mixed PSCs/BxPC-3 cells treated with follistatin (20 ng/ml) every 3 days and collected tumor samples at day 14 after inoculation. Representative IHC images t and H score u ofαSMA expression. Scale bar, 0.5 mm. Values were presented as mean ± SD (n = 5). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 (two-tailed t-test).

false

• WB

CiteAb

Western blot - Anti-Collagen I + Collagen III antibody (AB34710)

Western Blotting using Anti-Collagen I + Collagen III antibody, ab34710. Publication image from Yao, H. et al., 2015, Nat Commun, 26436920. Legend direct from paper.

miR-142-5p and miR-130a-3p regulate the profibrogenesis of macrophages.(a–g) Human macrophages were transduced with control, miR-142-5p ASO or miR-130a-3p mimics or both. After 24 h, the macrophages were stimulated with IL-4 for 12 h and co-cultured with primary human fibroblasts for another 48 h. (a) The schematics of the approach. (b) Representative images of fluorescent FAP/DAPI staining in fibroblasts. Scale bar, 100µm. Fluorescence intensity was quantitated.(mean±s.e.m., n=3 independent experiments; *P<0.05; **P<0.01. P values were obtained using two-tailed Student's t-test). (c) Fibroblast contractility in three-dimensional collagen matrices (mean±s.e.m., n=4 independent experiments; *P<0.05; **P<0.01. P values were obtained using two-tailed Student's t-test). (d) Representative images of western blot analysis ofα-SMA, collagen I and collagen III in fibroblasts (n=3). The numbers above the blots present the intensity ratio of indicated protein/GAPDH analysed by ImageJ. (e) Extracellular acid-soluble collagen production of fibroblasts was measured by the Sircol assay. (mean±s.e.m., n=4 independent experiments; *P<0.05; **P<0.01; ***P<0.001. P values were obtained using two-tailed Student's t-test). (f) The proliferation of fibroblasts was determined by BrdU incorporation assay (mean±s.e.m., n=4 independent experiments; *P<0.05; **P<0.01. P values were obtained using two-tailed Student's t-test). (g) Total TGF-β1 (acid-treated) and active TGF-β1 (not acid-treated) in the media of the macrophage/fibroblast co-culture system (mean±s.e.m., n=4 independent experiments; *P<0.05; **P<0.01; ***P<0.001. P values were obtained using two-tailed Student's t-test).

false

• WB

CiteAb

Western blot - Anti-Collagen I + Collagen III antibody (AB34710)

Western Blotting using Anti-Collagen I antibody, ab34710. Publication image from Jeng, Y. M. et al., 2022, Nat Commun, 35618735. Legend direct from paper.

Direct tumor-fibroblast contact triggers activin A secretion leading to EMT of tumor cells and myofibroblast activation.a Human cytokine array used conditional medium from transwell (indirect, ID) versus direct (D) co-culture of BxPC-3 with pancreatic stellate cells (PSCs). Cytokine levels in direct co-cultured condition were normalized to indirect co-cultured condition. b Quantitative real-time PCR analyses of CLDN1, VIM, Snail, Twist1, ZEB1, and ZEB2 expression were performed in BxPC-3 cells treated with recombinant human Activin A (50 and 100 ng/ml) compared to untreated cells. Values were presented as mean ± SD (n = 3). c Western blotting analysis of proteins harvested from BxPC-3 cells treated with recombinant human activin A (50 and 100 ng/ml) compared to untreated cells. d Sphere-formation assay of BxPC-3 cells treated with recombinant human activin A compared to untreated cells. Values were presented as mean ± SD (n = 4). e Western blotting analysis of proteins harvested from mono-cultured and direct co-cultured (D) tumor cells (BxPC-3-RFP) treated with IgG (1 µg/ml) or anti-activin A-neutralizing antibody (1 µg/ml, 693604, Biolegend). FACS was performed to collect RFP+ tumor cells after 48 h of mono- or co-culture conditions. f Sphere-formation assay of BxPC-3-RFP cells in mono-cultured and direct co-cultured conditions treated with IgG (2 µg/ml) or anti-activin A-neutralizing antibody (1 and 2 µg/ml). Values were presented as mean ± SD (n = 4). g, h 3D spheroid invasion assay of BxPC-3-RFP spheroids in mono-cultured and direct co-cultured conditions treated with IgG (4 µg/ml) or anti-activin A-neutralizing antibody (2 and 4 µg/ml). g Representative images. Scale bar, 200 µm. h Extensive rate of each tumor spheroid was calculated by normalizing RFP+ area at 72 h to RFP+ area at 1 h (left panel) and by normalizing RFP+ maximal diameter at 72 h to RFP+ maximal diameter at 1 h (right panel). Values were presented as mean ± SD (n = 9). i, j Subcutaneous injection of BxPC-3 cells and mixed fibroblasts/BxPC-3 cells treated with IgG (1 µg/ml) or anti-activin A-neutralizing antibody (1 µg/ml) every 3 days and collected tumor samples at day 14 after inoculation. Representative images i and quantification results of tumor weight j. Scale bar, 1 cm. Values were presented as mean ± SD (n = 6). k, l Subcutaneous injection of BxPC-3 cells and mixed fibroblasts/BxPC-3 cells treated with recombinant human follistatin 20 ng/ml every 3 days and collected tumor samples at day 14 after inoculation. Representative images k and quantification results of tumor weight l. Scale bar, 1 cm. Values were presented as mean ± SD (n = 5). m, n Quantitative real-time PCR analyses and western blotting analysis of ACTA2 and COL1A1 expression were performed in PSCs treated with recombinant human activin A (50 and 100 ng/ml) compared to untreated cells for 24 h and 48 h, respectively. For qPCR analyses, values were presented as mean ± SD (n = 3). o, p Quantitative real-time PCR analyses and western blotting analysis of ACTA2 and COL1A1 expression in mono-cultured, indirect co-cultured (ID), and direct co-cultured (D) fibroblasts (PSC-GFP) with tumor cells (BxPC-3). FACS was performed to collect GFP+ fibroblasts after 24 h and 48 h for mRNA and protein samples, respectively. For qPCR analyses, values were presented as mean ± SD (n = 3). q Western blotting analysis of proteins harvested from mono-cultured and direct co-cultured (D) fibroblasts (PSC-GFP) treated with IgG (2 µg/ml) or anti-activin A-neutralizing antibody (1 and 2 µg/ml). FACS was performed to collect GFP+ fibroblasts after 48 h of mono- or co-cultured conditions. r, s BxPC-3 cells and mixed PSCs/BxPC-3 cells treated with IgG (2 µg/ml) or anti-activin A-neutralizing antibody (2 µg/ml) every 3 days and collected tumor samples at day 14 after inoculation. Representative IHC images r and H Score s ofαSMA expression. Scale bar, 0.5 mm. Values were presented as mean ± SD (n = 6). t, u BxPC-3 cells and mixed PSCs/BxPC-3 cells treated with follistatin (20 ng/ml) every 3 days and collected tumor samples at day 14 after inoculation. Representative IHC images t and H score u ofαSMA expression. Scale bar, 0.5 mm. Values were presented as mean ± SD (n = 5). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001 (two-tailed t-test).

false