Cd3 percp cyanine5

Alginate sodium (160 mPa·s viscosity) with low G/M ratio was purchased from Qingdao Crystal Salt Bioscience and Technology Corporation (China). The G/M ratio of the alginate sodium was 1:3, as determined by circular dichroism (CD) spectrometer.19 (link) Endostar was bought from Shandong Xiansheng Maidejin Biological Pharmaceutical Co., Ltd (China). Dopamine hydrochloride was purchased from Yuancheng Technology Development Co., Ltd. (China). Mouse VEGF, matrix metalloproteinase-9 (MMP-9), IL-8 and IL-17 ELISA kits were obtained from Shanghai Enzyme-Linked Biotechnology Co., Ltd. (China). Mouse IFN‑γ, TNF-α, IL-4, IL-6, IL-10 and IL-12 ELISA kits, and CD11c-PerCP-Cyanine5.5, MHCI-APC and CD86-PE were the products of eBioscience (USA). Mouse monoclonal antibodies MHCII-FITC, CD3-PerCP/Cyanine5.5, CD8-APC and CD4-FITC were purchased from BioLegend (USA). MTT Cell Proliferation and Cytotoxicity Assay Kit, penicillin, streptomycin and goat serum were purchased from Beyotime (China). In situ Cell Death Detection Kit (POD), Drabkin’s reagent, and hemoglobin standard were obtained from Sigma-Aldrich, Inc. (USA). RPMI-1640 medium and heat-inactivated FBS were purchased from Biological Industries (Israel).

Mouse skin tissues were cut into small pieces and digested for 2 hours in RPMI1640 medium containing 1% FBS, 1 mg/mL collagenase type 1A, 50 µg/mL DNaseI and 400 µg/mL Hyaluronidase (all from Sigma) at 37°C with rotation, and then filtered through 40 mm cell strainers. Cervical lymph nodes and spleens from mice were produced by gentle grinding with a 200-mesh iron sand mesh. Red blood cells were removed using ACK Lysis Buffer (Beyotime, China). For the analysis of IL-17A production, mouse antibodies including: Zombie Aqua™ dye-live/dead, CD3-PerCP/Cyanine5.5, γδTCR-PE, CD4-FITC, IL17A-PE/Cyanine7, were purchased from Biolegend. Samples were harvested with BD FACS Canto and analyzed with FlowJo software.