Biglycan

Manufactured by R&D Systems

Sourced in United States

Biglycan is an extracellular matrix protein that belongs to the small leucine-rich proteoglycan (SLRP) family. It is involved in the regulation of collagen fibrillogenesis and the assembly of the extracellular matrix.

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Lab products found in correlation

Fibronectin, by R&D Systems (1 mentions) Aggrecan, by R&D Systems (1 mentions) Matrigel, by R&D Systems (1 mentions) Matrigel, by BD (1 mentions) Enhanced chemiluminescence, by Thermo Fisher Scientific (1 mentions) Protease and phosphatase inhibitor cocktail, by Merck Group (1 mentions) β actin, by Thermo Fisher Scientific (1 mentions) β tubulin, by Santa Cruz Biotechnology (1 mentions) Snx 482, by Bio-Techne (1 mentions) Hmbg1, by R&D Systems (1 mentions) Tenascin c, by R&D Systems (1 mentions) Adenosine triphosphate (atp), by Merck Group (1 mentions) Pam2csk4, by Thermo Fisher Scientific (1 mentions)

5 protocols using biglycan

Optimizing Stem Cell Culture Conditions

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The following soluble factors at designated concentrations were used in the study: bone morphogenetic factor 6 (BMP6, R&D systems, 100 ng/ml), 1α,25-dihydroxyvitamin D3 (VD3, Sigma Aldrich, 100 nM), valproic acid (VPA, Sigma Aldrich, 2 mM), Wnt3a (R&D systems, 100 ng/ml), Wnt5a (R&D systems, 100 ng/ml), and Dickkopf1 (Dkk1, R&D systems, 100 ng/ml).
Among ECM components, we used aggrecan, biglycan, fibronectin, matrigel, collagen type I, hyaluronic acid (HA), and chondroitin sulphate A. Multiwell plates were coated with aggrecan (R&D systems) at 0.4 nmol/cm², biglycan (R&D systems) at 0.2 nmol/cm², fibronectin (R&D systems) at 0.4 nmol/cm², chondroitin sulphate A (CS) (Sigma Aldrich) at 0.05 mg/cm², or 0.05 mg/cm² HA (Abcam, MW 1.6 kDa). Type I collagen solution extracted from the rat tail tendons was incubated at 37°C for 30 min. matrigel (BD Biosciences) was diluted at a concentration 0.12 mg/cm² and incubated at 4°С overnight.

Kalabusheva E., Terskikh V, & Vorotelyak E. (2017). Hair Germ Model In Vitro via Human Postnatal Keratinocyte-Dermal Papilla Interactions: Impact of Hyaluronic Acid. Stem Cells International, 2017, 9271869.

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Quantifying Atherosclerosis in Mouse Aorta

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Atherosclerosis was quantified in the aortic sinus and the aortic intimal surface as described previously[9 (link)]. Aortic sinus sections (10 μm thick) collected every 90 μm were stained with Oil Red O (Fluka, St. Louis, MO, USA) and quantified using computer-assisted morphometry with Nikon Image software. Aortic intimal surface lesions were stained with Sudan IV (MP Biomedicals, Solon, OH, USA) then evaluated by en-face quantification of lesions. All quantifications were performed in duplicate by observers blinded to the group assignments of the mice. Immunofluorescence staining was performed using primary antibodies against biglycan (R&D Systems, Minneapolis, MN, USA), perlecan (Accurate, Westbury, NY, USA), or apoB (Biodesign International, Saco, ME, USA) and analyzed by confocal microscopy with the use of a Leica AOBS TCS SP5 inverted laser scanning confocal microscope (Leica Microsystems Inc. Mannheim, Germany). Autofluorescence was eliminated by Sudan Black treatment as previously described[13 ]. Negative controls were performed using isotype-matched irrelevant primary antibodies, no primary antibody, or no secondary antibody. Perlecan immunostaining was performed on aortic cross sections and visualized by light microscopy and quantified as % aortic area using ImageJ software as previously described[13 ].

Tang T., Thompson J.C., Wilson P.G., Yoder M.H., Müeller J., Fischer J.W., Jon Williams K, & Tannock L.R. (2014). Biglycan deficiency: increased aortic aneurysm formation and lack of atheroprotection. Journal of molecular and cellular cardiology, 75, 174-180.

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Protein Extraction for Western Blot Analysis

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To extract the protein for western blot analysis, cells were harvested by scraping on ice with 1× SDS-sample buffer containing protease and phosphatase inhibitor cocktails (Sigma-Aldrich, St. Louis, MO, USA) and then heated at 95 °C for 5 min. In addition, conditioned media were collected to detect secreted proteins. An equivalent volume of conditioned media from an equal number of cells were mixed with 4× SDS-sample buffer, and heated at 95 °C for 5 min. Equal amounts of samples were loaded and separated by SDS-PAGE and transferred onto nitrocellulose membranes. Following blocking with 5% skim milk, membranes were immunoblotted with primary anti-human decorin, biglycan (R&D Systems Inc., Minneapolis, MN, USA), β-actin (Thermo Fisher Scientific), β-tubulin (Santa Cruz Biotechnology, Santa Cruz, CA, USA), ADAMTS5 (GeneTex, Irvine, CA, USA), or monoclonal anti-procollagen type I amino-terminal extension peptide (SP1.D8) (Developmental Studies Hybridoma Bank, Iowa City, IA, USA) antibody, and polyclonal (GeneTex) or monoclonal secondary antibody (Santa Cruz Biotechnology), then detected using enhanced chemiluminescence (Thermo Fisher Scientific).

Lee H., Lim J., Oh J.H., Cho S, & Chung J.H. (2021). IGF-1 Upregulates Biglycan and Decorin by Increasing Translation and Reducing ADAMTS5 Expression. International Journal of Molecular Sciences, 22(3), 1403.

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Monocyte/Macrophage Activation by DAMPs

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The selection of DAMPs was done according to the literature [e.g., ref. (13 (link)–18 (link))] and based on following criteria: (1) known receptors, (2) receptors present on monocytes/macrophages, (3) described effect on monocyte/macrophages, (4) released as alarmins from other cells/tissues, and (5) relevance to RA.
Recombinant human proteins were used: HSP90/GRP94 (Abcam), tenascin C, HMBG1, biglycan, and S100A4 (R&D Systems). In addition, purified human oS100A4 was provided by M. Grigorian (Copenhagen University, Copenhagen, DK). β-Glucan isolated from Candida albicans was provided by the department of internal Medicine, Radboudumc (Nijmegen, NL). Other compounds used in the current study included human fibrinogen and citrullinated fibrinogen (Cayman), muramyl dipeptide (MDP), E. coli lipolpolysaccharide-B5, Salmonella typhimurium flagellin and ATP (Sigma). All DAMPs were tested for endotoxin contamination, using the Pierce LAL Chromogenic Endotoxin Quantification Kit. SNX482 (Tocris) was diluted to 25 nM in RPMI.

Neidhart M., Pajak A., Laskari K., Riksen N.P., Joosten L.A., Netea M.G., Lutgens E., Stroes E.S., Ciurea A., Distler O., Grigorian M, & Karouzakis E. (2019). Oligomeric S100A4 Is Associated With Monocyte Innate Immune Memory and Bypass of Tolerance to Subsequent Stimulation With Lipopolysaccharides. Frontiers in Immunology, 10, 791.

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Chondrocyte Inflammatory Response Assay

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Monolayer chondrocytes were used at passage 2. 150 000 cells were seeded in 6 well plates and were cultured in complete chondrocyte media to confluent. At confluence, cells were pre‐treated with 1/1000 dilution of Sparstolonin B, o‐Vanillin or PBS in serum‐free chondrocyte media for 1 hour before adding a final concentration of 1 μg/mL S100A8/9 (R&D Systems) or 2.5 µg/mL biglycan (R&D Systems). The synthetic agonist Pam2CSK4 (Invitrogen) was also used here as a positive control at a lower concentration of 1 ng/mL. The lower dose was chosen for physiological relevance to simulate the low‐grade inflammation found in OA. For gene expression analysis, cells were lysed in TRIzol 16 hours following alarmin treatment. Culture media were collected after 48 hours.

Bisson D.G., Sheng K., Kocabas S., Krock E., Teles A., Saran N., Ouellet J.A, & Haglund L. (2020). Toll‐like receptor involvement in adolescent scoliotic facet joint degeneration. Journal of Cellular and Molecular Medicine, 24(19), 11355-11365.

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