Human total mmp 1 elisa kit

Using a fluorescence microplate reader (RF-5301PC; Shimadzu Corp., Tokyo, Japan) and adapting Losso et al.’s method [36 (link)], HDF cells were cultured (2 × 10⁴ cells/well) for 24 h. Following this, cells were exposed to KO (0.25, 0.5, 1, 1.5, 2, 4, and 8 mg/mL) or 1 μM RA, with or without 5 mJ/cm² UVB irradiation for 2 min and cultured for an additional 24 h. Total MMP-1 protein levels in supernatants were quantified using the Human Total MMP-1 ELISA kit (DY901; R&D Systems, Minneapolis, MN, USA), normalized to total protein content. Relative MMP-1 expressions (%) were calculated as [(MMP-1 contents in UVB-exposed control or experimental sample/MMP-1 contents in unexposed control) × 100]. IC₅₀ values were reported as concentrations where UVB-induced HDF cell MMP-1 expressions were 50% inhibited. UVB irradiation was performed using a UV irradiation Crosslinker system (CL-1000M, Analytik Jena, Upland, CA, USA).

The concentration of MMP1 in supernatants of synovial fibroblasts transfected with HOTAIR siRNA or AllStars Negative siRNA control was measured using the human total MMP1 ELISA kit (R&D Systems, according to the manufacturer's instructions) and the GloMax-Multi+Detection System (Promega) with Instinct Software (Promega).

Cultured cell supernatants were prepared and analyzed using quantikine human TIMP‐1 ELISA kit, human TGF‐β1 Duoset ELISA kit, human total MMP‐1 ELISA kit, and human MMP‐9 ELISA kit (R&D Systems, Minneapolis, MN, USA). ELISAs were conducted according to the manufacturer's instructions. All samples were analyzed in triplicates in each experiments.