Blyscan sgag assay

Manufactured by Biocolor

Sourced in United Kingdom

The Blyscan sGAG assay is a biochemical test used to quantify the levels of sulfated glycosaminoglycans (sGAGs) in biological samples. The assay provides a sensitive and reproducible method for measuring sGAG content, which is important in various areas of research and clinical applications.

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Lab products found in correlation

Dneasy blood tissue kit, by Qiagen (1 mentions) Calf thymus dna, by Merck Group (1 mentions) Hoechst 33 258, by Biosharp (1 mentions) Cysteine hcl, by Merck Group (1 mentions) Papain, by Merck Group (1 mentions) Sodium acetate, by Merck Group (1 mentions) Quant it picogreen dsdna assay, by Thermo Fisher Scientific (1 mentions) Picogreen dsdna assay, by Thermo Fisher Scientific (1 mentions) Heto powerdry ll1500 freeze dryer, by Thermo Fisher Scientific (1 mentions)

Close spelling variants of this product

Blyscan sGAG assay kit Blyscan sulfated glycosaminoglycan (sGAG) assay kit Blyscan™ sGAG Assay kit (B1000, Blyscan sulfated sGAG assay kit Blyscan assay Blyscan

7 protocols using blyscan sgag assay

Quantification of Cartilage sGAG Content

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In this study, sGAG content in each specimen was quantified using
the Blyscan sGAG Assay (Biocolor). The assay was performed following
the manufacturer’s protocol. In summary, a full-thickness piece
of cartilage was obtained from three pairs (a total of six pinnas,
three from each study group). The auricle pairs were randomly selected
from the total pairs included in the study. The weight of each cartilage
specimen was recorded. The sGAG was extracted from the tissue using
a Papain extraction reagent. A dye was then added to form a complex
with sGAG. The dye-bound sGAG was precipitated and drained. Subsequently,
a dissociation agent was added to unbind the sGAG from the dye. The
recovered sGAG content was measured at 656 nm using spectrophotometry
(SpectraMax Plus 384 microplate reader) and estimated using a standardized
curve.

Al-Qurayshi Z., Wafa E.I., Rossi Meyer M.K., Owen S, & Salem A.K. (2021). Tissue Engineering the Pinna: Comparison and Characterization of Human Decellularized Auricular Biological Scaffolds. ACS Applied Bio Materials, 4(9), 7234-7242.

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Quantification of Laryngeal sGAG

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sGAG content of cricothyroid muscle and cricoid cartilage from three randomly selected larynges was quantified using Blyscan^™ sGAG Assay (Biocolor, Northern Irland, UK). This assay is widely used to measure sGAG content in cartilages, organs/tissues (such as heart, lungs, kidney, larynx, and trachea), and solid tumors.^{12 (link)–14 (link)} The assay was performed following the manufacturer’s protocol. In summary, the weight of each of the muscle and cartilage specimens (3 each) was recorded. The sGAG was extracted from the tissue using a Papain extraction reagent. Then, a dye was added that forms a complex with sGAG. The dye-bound sGAG was precipitated and drained. Subsequently, a dissociation agent was added to unbind the sGAG from the dye. The recovered sGAG content was measured at 656 nm using spectrophotometry and estimated by plotting on a standardized curve.

Al-Qurayshi Z., Wafa E.I., Hoffman H., Chang K, & Salem A.K. (2021). Tissue-engineering the Larynx: Effect of Decellularization on Human Laryngeal Framework and the Cricoarytenoid Joint. Journal of biomedical materials research. Part B, Applied biomaterials, 109(12), 2030-2040.

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Extracellular Matrix Composition Analysis

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Collagen content was determined using a hydroxyproline assay (QuickZyme) according to manufacturer’s instructions (n=7). Sulfated glycosaminoglycan (sGAG) content was determined using the Blyscan sGAG assay (Biocolor). Samples were digested in papain overnight at 65°C overnight before proceeding with protocol according to manufacturer’s instructions (n=3). Residual DNA was quantified with a DNeasy Blood & Tissue Kit (Qiagen) according to manufacturer’s instructions (n=3).

Morris A.H., Stamer D.K., Kunkemoeller B., Chang J., Xing H, & Kyriakides T.R. (2018). Decellularized materials derived from TSP2-KO mice promote enhanced neovascularization and integration in diabetic wounds. Biomaterials, 169, 61-71.

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Quantitative Analysis of GAG and DNA

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At the specified time points, the cells were washed with PBS three times and then were frozen in liquid nitrogen and lyophilized for 12 h. The remaining cells were collected and digested with the GAG digestion solution (papain 0.0156 g, NaOH 0.2 g, EDTA 0.0731 g, sodium dihydrogen phosphate 0.39 g, and cysteine 0.0304 g in 25 ml ultrapure water) for 16 h at 60°C. The GAG content was measured using the Blyscan sGAG Assay (Biocolor, United Kingdom), and the standard curve was generated by chondroitin sulfate standard using the kit. The DNA content was quantified using Hoechst 33,258 (Biosharp, China). Calf thymus DNA (Sigma) was used to generate a standard curve (Farndale et al., 1986 (link)).

Yuan Z., Liu S., Song W., Liu Y., Bi G., Xie R, & Ren L. (2022). Galactose Enhances Chondrogenic Differentiation of ATDC5 and Cartilage Matrix Formation by Chondrocytes. Frontiers in Molecular Biosciences, 9, 850778.

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Quantification of Sulfated Glycosaminoglycans

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After completion of the crosslinking and decellularization process, three P1 scaffolds or one P14 ECM scaffold were flash frozen and lyophilized. The powder was solubilized and digested overnight at 65 °C (0.2 M Na₂HPO₄/NaH₂PO₄ buffer (pH 6.4), 0.01 M EDTA disodium salt dihydrate, 0.14 mg/mL papain (Sigma, St. Louis, MO, USA, P3125), 0.09 M sodium acetate (Merck, Rahway, NJ, USA, 6268) and 5 mM cysteine HCl (Sigma, St. Louis, MO, USA, 30120)) under gentle shaking. After centrifugation, 100 µL of supernatant was collected for the quantification of sulfated glycosaminoglycans using the Blyscan sGAG assay (Biocolor, Carrickfergus, Northern Ireland, B1000) according to the manufacturer’s instruction [45 (link)].

Glorieux L., Vandooren L., Derclaye S., Pyr dit Ruys S., Oncina-Gil P., Salowka A., Herinckx G., Aajja E., Lemoine P., Spourquet C., Lefort H., Henriet P., Tyteca D., Spagnoli F.M., Alsteens D., Vertommen D, & Pierreux C.E. (2023). In-Depth Analysis of the Pancreatic Extracellular Matrix during Development for Next-Generation Tissue Engineering. International Journal of Molecular Sciences, 24(12), 10268.

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Quantification of sGAG and DNA in PCL-CHyA

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CHyA scaffolds sGAG and DNA quantification were both performed by first lysing cell-seeded PCL-CHyA and CHyA using consecutive freeze-thaw cycles at -80 °C prior to papain (0.01% wt/wt) digestion (Sigma-Aldrich) overnight at 65 °C. Following incubation, the respective assay manufacturer's protocols were followed for the Blyscan sGAG assay (Biocolor, Carrickfergus, UK) (Matsiko et al., 2015) (link) and the Quant-iT PicoGreen dsDNA assay (Invitrogen). In brief, sGAG were quantified by binding with Blyscan dye and absorbance measured at a wavelength of 656 nm. sGAG concentration was determined by comparison to a standard curve. Similarly, DNA was quantified by intercalating dsDNA with Picogreen and measuring the fluorescence at a wavelength of 538 nm. Then, dsDNA concentration was determined by comparison to a standard curve.

Joyce M., Hodgkinson T., Lemoine M., González-Vázquez A., Kelly D.J, & O'Brien F.J. (2023). Development of a 3D-printed bioabsorbable composite scaffold with mechanical properties suitable for treating large, load-bearingarticular cartilage defects. European cells & materials, 45.

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Decellularization of Rat Kidney

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Approximately half of a whole, unfixed rat kidney or kidney bioscaffold was suspended within 5 ml of papain solution (55 mM L-cysteine hydrochloride, 80 mM EDTA, 2.5 units/ml papain, pH 6.0, all Sigma/UK) for 24 h at 60 o C until no visible solid material remained. After digestion, the solution was freeze-dried for 24h using the Heto PowerDry LL1500 freeze dryer (Thermo Scientific/UK) to ascertain the dry weight.
Total DNA content was quantified using the Picogreen® dsDNA assay (Invitrogen/UK) and total sGAG content was quantified using the Blyscan™ sGAG assay (Biocolor/UK) according to the manufacturer's instructions. Each assay was performed in duplicate and three times in total.

He M., Callanan A., Lagaras K., Steele J.A.M, & Stevens M.M. (2017). Optimization of SDS exposure on preservation of ECM characteristics in whole organ decellularization of rat kidneys. Journal of biomedical materials research. Part B, Applied biomaterials, 105(6).

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