Human bmp 2 elisa kit

To investigate the immunomodulatory effects of AMM/B on synovial fibroblast, 5 × 10⁵ synovial fibroblasts were treated without or with 10 ng/mL tumor necrosis factor (TNF)-α for 24 h and then co-cultured with 5 × 10⁵ AMMs or AMM/B in RPMI 1640 containing 10% FBS. The co-cultured supernatants were harvested after two days, and the IL34 cytokine levels were evaluated using a murine IL-34 ELISA kit (R&D Systems, Minneapolis, MN, USA). The concentration of IL-17A was measured using murine IL-17A ELISA kits (R&D Systems) and mouse serum. The BMP-2 protein level was also examined using a human BMP-2 ELISA kit (R&D Systems).

To quantify the BMP2 and VEGF-A expression following BMSC transfection, the supernatant was sampled at different days post-transfection (dpt) for ELISA analysis using the human BMP-2 ELISA kit (Quantikine, R&D Systems, USA) and human VEGF ELISA kit (Quantikine, R&D Systems, USA), according to the manufacturer’s instructions. The absorbance was measured at 450 nm. Wavelength correction was set at 570 nm. Experiments were performed in triplicates, and the protein content was determined using a standard curve (range: 0-1000 pg ml⁻¹ BMP-2 and 0-1000 pg ml⁻¹ VEGF).

Dulbecco’s modified eagle medium (DMEM) and fetal bovine serum (FBS) were purchased from Gibco (Thermo Fisher Scientific, USA). Trypsin-EDTA (0.25%, 1× solution), Dulbecco’s phosphate buffered saline (DPBS) and antibiotics (100 U mL⁻¹ penicillin G and 0.1 mg ml⁻¹ streptomycin) were purchased from Gibco® (Invitrogen™, Grand Island, NY). Cell Counting Kit-8 (CCK-8) was obtained from Dojindo Laboratories (Dojindo, Kumamoto, Japan). Alizarin Red S was purchased from Solarbio^®Life Sciences, Alkaline phosphatase (ALP) staining kit (BCIP/NBT Alkaline Phosphatase Color Development Kit) was purchased from Beyotime Institute of Biotechnology. Collagen was obtained from Sichuan Mingrang Tech., China and collagen cross-linking agent N-(3-(dimethylamino)-propyl)-N-Ethylcarbodiimide hydrochloride crystalline (EDC) and N-Hydroxysulfosuccinimide sodium salt (NHS) were purchased from Aladdin Industrial Corporation, Shanghai, China. Opti-MEM medium and Lipofectamine^® MessengerMAX^TM Reagent were obtained from Gibco^TM (Invitrogen, CA, USA). Human BMP-2 ELISA kit and human VEGF ELISA kit were obtained from R&D (Quantikine, R&D Systems, USA). D-Luciferin, Sodium Salt was purchased from Yeasen (Shanghai, China). All materials were utilized as received, without any further purification.