Human total mmp 1 duoset elisa

Heat-inactivated fetal bovine serum (FBS, HyCylone™) and PBS (10X), pH 7.4 were purchased from Thermo Fisher Scientific (Waltham, MA, USA). Dulbecco’s modified Eagle’s medium high glucose 4.5 g/L (DMEM), penicillin-streptomycin solution, trypsin-EDTA (0.25%), 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT), 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS substrate), 3,3′,5,5′-tetramethylbenzidine (TMB substrate), human glycated albumin, elastase from human leukocytes, N-succinyl-Ala-Ala-Ala-p-nitroanilide, oleanolic acid, hyaluronidase from bovine testes, 4 (dimethylamino)benzaldehyde (DMAB), hyaluronic acid sodium salt from cockscomb, sodium aurothiomalate hydrate (SATMH) and Collagenase Activity Colorimetric Assay Kit (MAK293) were purchased from Merck (Waltham, MA, USA). ELISA Human IL-6 Kit (900-K16) and ELISA Human IL-8 Kit (900-K18) were purchased from PeproTech (London, UK). Human Total MMP-1 DuoSet ELISA (DY901B-05), Human MMP-2 Duo-Set ELISA (DY902), and Human Pro-Collagen I alpha 1 DuoSet ELISA (DY6220-05) were purchased from R&D Systems (Minneapolis, MI, USA).

To measure MMP-1 and type I procollagen, the HDFs were plated in 24-well plates at a density of 1 × 10⁵ cells/well. Cells were treated with or without DEQA (final concentrations of 1, 5 and 20 μM) immediately after UVA irradiation (10 J/cm²). After 24 h, the supernatants were collected and quantified for MMP-1 and type I procollagen contents by using Human Total MMP-1 DuoSet ELISA (#DY901B; R&D System Inc., Minneapolis, MN, USA) and Human Pro-Collagen I alpha 1 DuoSet ELISA (#DY6220; R&D System Inc., Minneapolis, MN, USA) kits, respectively, according to manufacturer’s instructions.