Satellite cells were isolated as described above and seeded in plates coated with collagen I (1 μg mL−1, BD Bioscience) and laminin (10 μg mL−1, BD Bioscience) in myogenic growth media (20% heat-inactivated horse serum, 1% pencillin/streptomycin and 5 ng mL−1 bFGF in F10). 384-well plates were used for all EdU proliferation assays using ~2,000 cells per well. 24-well plates were used for signaling experiments using ~150,000 cells. Cells were treated with recombinant plasminogen (Abcam) at 5 μg/ml or 54 nmol/L or plasmin (Abcam) at 5 μg/ml or 57 nmol/L. Additional groups were treated with the ERK inhibitor LY3214996 (MedChemExpress) at 2.3 μg/ml or 5 μmol/L.
Plasmin
Manufactured by Abcam
Plasmin is a serine protease enzyme that plays a crucial role in the dissolution of fibrin clots. It is the active enzyme form of the precursor plasminogen and is involved in the breakdown of extracellular matrix proteins.
Automatically generated - may contain errors
Lab products found in correlation
Laminin, by BD (1 mentions)
Ly3214996, by MedChemExpress (1 mentions)
Collagen 1, by BD (1 mentions)
Imidazole, by Merck Group (1 mentions)
Sodium deoxycholate, by Merck Group (1 mentions)
Sequencing grade trypsin, by Promega (1 mentions)
Tris hcl, by Merck Group (1 mentions)
Tris 2 carboxyethyl phosphine, by Merck Group (1 mentions)
Oasis hlb 96 well μelution plate, by Waters Corporation (1 mentions)
Cobalt loaded resin, by Thermo Fisher Scientific (1 mentions)
Imperial protein stain, by Bio-Rad (1 mentions)
Sds page standard, by Bio-Rad (1 mentions)
Chloroacetamide, by Merck Group (1 mentions)
Acetonitrile, by Biosolve (1 mentions)
Formic acid, by Biosolve (1 mentions)
2 protocols using plasmin
1
Satellite Cell Proliferation Assay
2
Targeted HRG Variant Production
Check if the same lab product or an alternative is used in the 5 most similar protocols
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Human plasma containing EDTA from a single healthy individual was obtained from Zen-Bio. One recombinant HRG variant, containing the mutations Pro204 and Asn493, was produced in CHO cell lines by PeproTech. Four other HRG variants, with pairwise combinations of targeted mutations for positions 204 and 493, were produced in HEK293E-253 cells (HEK293) by ImmunoPrecise. HRGs were expressed with sequences matching allelic variants on position 204 or 493 in human population: Pro204/Asn493, Pro204/Ile493, Ser204/Ile493, and Ser204/Asn493.
Cobalt-loaded resin was acquired from Thermo Scientific. Imidazole was purchased from Sigma–Aldrich. Tris–HCl, Tris(2-carboxyethyl)phosphine, chloroacetamide, sodium deoxycholate (SDC), and TFA were purchased from Sigma–Aldrich. Oasis HLB 96-well μElution Plates were purchased from Waters. Sequencing-grade trypsin was obtained from Promega. SialEXO and OpeRATOR were obtained from Genovis. Plasmin was purchased from Abcam. Formic acid (FA) and acetonitrile were acquired from Biosolve Chimie. The SDS-PAGE standards and Imperial Protein Stain were purchased from Bio-Rad and Thermo Scientific, respectively.
Cobalt-loaded resin was acquired from Thermo Scientific. Imidazole was purchased from Sigma–Aldrich. Tris–HCl, Tris(2-carboxyethyl)phosphine, chloroacetamide, sodium deoxycholate (SDC), and TFA were purchased from Sigma–Aldrich. Oasis HLB 96-well μElution Plates were purchased from Waters. Sequencing-grade trypsin was obtained from Promega. SialEXO and OpeRATOR were obtained from Genovis. Plasmin was purchased from Abcam. Formic acid (FA) and acetonitrile were acquired from Biosolve Chimie. The SDS-PAGE standards and Imperial Protein Stain were purchased from Bio-Rad and Thermo Scientific, respectively.
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