Rabbit anti collagen 1

Manufactured by Novus Biologicals

Sourced in United States

Rabbit anti-collagen-1 is a primary antibody that recognizes collagen type I. It is used in various research applications to detect and study collagen I in biological samples.

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Close spelling variants of this product

Collagen I (21 citations) Anti‐Collagen I (6 citations)

5 protocols using rabbit anti collagen 1

Quantitative Analysis of Cell Secretome

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At 7 and 14 DIV, cells and conditioned media were harvested into SDS 1% solutions and subjected to a 5–20% gradient (cells lysates) or 7.5% (cell media) SDS-PAGE and to western blot analyses (as previously described57 (link)). Previous to immunoblot, ponceau S reversible staining was first used for detection of total protein content on nitrocellulose membranes58 (link). After overnight incubation at 4 °C with the primary antibodies (rabbit anti-collagen-I (1:1000); rabbit anti-osteonectin (1:500); mouse anti-β-Actin (1:1000), all from Novus Biologicals, Germany), horseradish peroxidase-linked (GE Healthcare, Chalfont St. Giles, UK) antibodies for enhanced chemiluminescence (ECL) detection were incubated 2 h/RT. Protein bands were scanned and quantified (GS-800™ Calibrated Densitometer and Quantity One densitometry software, Bio-Rad), and immunoblot data corrected to the respective ponceau loading control (as previously reported58 (link)).

Soares dos Santos M.P., Marote A., Santos T., Torrão J., Ramos A., Simões J.A., da Cruz e Silva O.A., Furlani E.P., Vieira S.I, & Ferreira J.A. (2016). New cosurface capacitive stimulators for the development of active osseointegrative implantable devices. Scientific Reports, 6, 30231.

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Comprehensive Lung Tissue Analysis

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The mice were euthanised and the lungs harvested and fixed with 4% paraformaldehyde diluted in PBS. Paraffin sections (2 μm) were stained with haematoxylin–eosin (H&E), Masson's trichrome and the following primary antibodies for immunohistochemistry: rabbit anti-E-cadherin, rabbit anti-α SMA, rabbit anti-vimentin, rabbit anti-MMP7 (Cell Signalling Technology, Danvers, MA, USA), rabbit anti-SP-C (Hycult Biotech, Uden, Netherlands), rat anti-Podoplanin (MBL, Aichi, Japan), rabbit anti-collagen I (Novus Biologicals, Littleton, CO, USA), rat anti-F4/80 (Bio-Rad Laboratories, Hercules, CA, USA), rabbit anti-CD3 (Genemed Biotechnologies, San Franciso, CA, USA), rat anti-PTPRC/CD45R (Aviva Systems Biology, San Diego, CA, USA), rabbit anti-arginase I (GeneTex, Irvine, CA, USA), mouse anti-peptidyl-citrulline (F95) and rabbit anti-S100A4 (Millipore, Burlington, MA, USA) and mouse anti-Laminin γ2 N-terminal fragment (γ2pf; Funakoshi, Tokyo, Japan). Histofine Simple Stain Mouse MAX-PO secondary antibodies (Nichirei, Tokyo, Japan) and the Opal 4-colour Fluorescent IHC kit (PerkinElmer, Waltham, MA, USA) were used according to the manufacturer's protocol. All images were captured by fluorescence microscopy (BZ-X710, Keyence) and analysed by hybrid cell count (Keyence, Osaka, Japan) or Fiji.

Miura Y., Ohkubo H., Niimi A, & Kanazawa S. (2021). Suppression of epithelial abnormalities by nintedanib in induced-rheumatoid arthritis-associated interstitial lung disease mouse model. ERJ Open Research, 7(4), 00345-2021.

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Renal Protein Extraction and Western Blot

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Renal tissue and cells were harvested and lysed by RIPA lysis buffer (Wuhan Goodbio Technology, China) containing cocktail protease inhibitors (Wuhan Goodbio Technology, China) for 30 min on ice. The total protein was obtained by high-speed centrifugation at low temperatures. Protein concentrations were quantified by a BCA protein assay kit (Beyotime Institute of Biotechnology, China), and 30 µg protein was loaded, separated on 10% or 12% SDS-PAGE, and transferred to nitrocellulose membranes. Then membranes were blotted at 4 °C overnight with mouse anti-VEGF-C (Santa Cruz, USA; 1:250), rabbit anti-Prox-1 (Angiobio, USA; 1:1000), rabbit anti-LYVE-1 (Novus, USA; 1:1000), Hamster anti-Podoplanin (Angiobio, USA; 1:200), rabbit anti-Collagen1 (Novus, USA; 1:2000), rabbit anti-α-SMA (Abcam, USA; 1:4000), rabbit anti-PDGFR-β (Abcam, USA; 1:2000), mouse anti-iNOS (Santa Cruz, USA; 1:200), rabbit anti-Arginase (Santa Cruz, USA; 1:400), rabbit anti-LC3B (Sigma-Aldrich, USA; 1:1000), rabbit anti-p62 (Abcam, USA; 1:5000), mouse anti-GAPDH (Wuhan Goodbio Technology, China; 1:2000), then were incubated with HRP-conjugated anti-IgG (Jackson ImmunoResearch, USA; 1:4000), finally were detected by ECL (Pierce, USA). Image capture and analysis were conducted by Bio-RAD (USA).

Zhang Y., Zhang C., Li L., Liang X., Cheng P., Li Q., Chang X., Wang K., Huang S., Li Y., Liu Y, & Xu G. (2021). Lymphangiogenesis in renal fibrosis arises from macrophages via VEGF-C/VEGFR3-dependent autophagy and polarization. Cell Death & Disease, 12(1), 109.

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Immunofluorescence Analysis of ONH LC Cells

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Primary human ONH LC cells were seeded on 12-well plates on coverslips and grown to confluency. After undergoing TLR4 inhibition and activation treatments as previously described for 72 hours, cells were washed with 1X PBS, fixed with 4% paraformaldehyde (PFA), permeabilized with 0.95% Triton X-100 in PBS, and blocked using Superblock Blocking Buffer in PBS (REF37580, Thermo Fisher Scientific) for 60 minutes at room temperature. Cells were labeled overnight at 4°C with rabbit anti-Fibronectin (F3648, Sigma-Aldrich) at a 1:100 dilution, or rabbit anti-collagen-1 (NB600–408, Novus Biologicals) at a 1:100 dilution in Superblock Blocking Buffer in PBS. Treatment without the primary antibody was used as a negative control. Coverslips were then incubated for at room temperature 2 hours using Alexa Fluor 488 donkey anti-rabbit IgG (REFA21206, Invitrogen – Thermo Fisher Scientific) at a 1:200 dilution. Coverslips were mounted to slides with Prolong Gold mounting medium containing DAPI (Invitrogen-Molecular Probes). Image acquisition was performed using Zeiss Axio Imager Z2 microscope. Images were taken at X20 or X40 magnification; scale bar represents 20μm.

Geiduschek E.K., Milne P.D., Mzyk P., Mavlyutov T.A, & McDowell C.M. (2022). TLR4 signaling modulates extracellular matrix production in the lamina cribrosa. Frontiers in ophthalmology, 2, 968381.

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Early TM Changes in Transgenic Mice

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To evaluate early changes in the TM, an initial cohort of mice were transduced with viral vectors, Ad5.Null (n = 7), Ad5.Cre (n = 15), and Ad5.TGFβ2 (n = 9), and harvested 11-days after injection. IOP was recorded at 10-days postinjection. Eyes were fixed in 4% PFA overnight, embedded in paraffin, cut into 5 μm sections, and transferred to glass slides. Deparaffinization was performed by washing two times with xylene, 100% ethanol, 95% ethanol, and 50% ethanol for 2 minutes each. Slides then were soaked in PBS for 5 minutes. Tissues were blocked using Superblock Blocking Buffer in PBS (Thermo Fisher Scientific) for 60 minutes. Rabbit anti-fibronectin (EMD Millipore) 1:500 dilution, rabbit anti-collagen-1 (Novus Biologicals, LL) 1:250 dilution, mouse anti-collagen-4 (Sigma-Aldrich Corp., St. Louis, MO, USA) 1:250 dilution, and mouse anti-BAMBI (Abnova; Walnut, CA, USA) 1:250 dilution, were used as primary antibodies, followed by Alexa-Fluor–labeled anti-rabbit or anti-mouse antibodies (Life Technologies, Carlsbad, CA, USA) 1:500 dilution. Prolong Gold mounting medium containing DAPI (Invitrogen-Molecular Probes) was used to mount the slides, and sections were imaged using the Keyence BZ-X700 fluorescence microscope (Keyence Corporation of America, Itasca, IL). All images were taken at ×100 magnification; scale bars represent 100 μm.

Hernandez H., Millar J.C., Curry S.M., Clark A.F, & McDowell C.M. (2018). BMP and Activin Membrane Bound Inhibitor Regulates the Extracellular Matrix in the Trabecular Meshwork. Investigative Ophthalmology & Visual Science, 59(5), 2154-2166.

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