Plusone coomassie tablets phastgel blue r 350

PlusOne Coomassie tablets PhastGel Blue R-350 and 12.5% ExcelGel were purchased from GE Healthcare Life Sciences (Uppsala, Sweden). CPDA1 blood bags were purchased from Medharmony (Qingdao, China). Dithiothreitol was purchased from Bio-Rad (Hercules, USA). Trypsin gold was purchased from Promega (Fitchburg, USA). Complete Protease Inhibitor Cocktail was purchased from Roche (Mannheim, Germany). Other chemicals and reagents were purchased from Sigma (Germany) unless indicated otherwise.

Supernatants from treated hDPCs were collected and quantified by using the Qubit® Protein Assay kit (Invitrogen). Samples were mixed with a nonreducing sample buffer (0.05 M Tris-HCL, pH 6.8, 2% sodium dodecyl sulphate [SDS], and 5% glycerol) and electrophoresed (150 V, 4°C) on 8% SDS-polyacrylamide gel with 1% gelatin from porcine skin, type A (Sigma-Aldrich). Afterwards, gels were rinsed twice with 2.5% Triton X-100 for 30 minutes at RT. They were then incubated overnight (37°C) in a reaction buffer (50 mM Tris-HCL, pH 7.4, and 10 mM CaCl₂), rinsed with distilled water, stained with 0.1% Coomassie blue (PlusOne Coomassie Tablets PhastGel® Blue R350, GE Healthcare, Chicago, IL) in 30% methanol/10% acetic acid solution for 30 minutes, and finally destained with 20% acetic acid solution. Band density was measured using ImageJ software (Rasband Wayne, National Institute of Health, Bethesda, MD). The molecular weight was estimated with Precision Plus Protein® Kaleidoscope® Standards (Bio-Rad, Hercules, CA).