Mouse il 17 elisa kit

Splenocytes isolated from BALB/c mice were stimulated with anti-CD3 or anti-CD3 + IL-6 + TGF-β with or without 1.25–10 nM triptolide treatment for 4 days (primary culture). The supernatants were assayed for IL-17 using a mouse IL-17 ELISA kit (ab100702, Abcam).
Serum concentrations of IL-17 and interferon (IFN)-γ were detected using the mouse IL-17 ELISA kit (ab100702, Abcam) and a mouse IFN-γ ELISA kit (ab252363, Abcam), respectively.

Two months after immunization and 40-45 days after injection of transduced cells, spleen cells were collected, cell suspensions were prepared in 96-well plates, and 50 ng/ml antigen (MOG35- 55 peptide) was added to RPMI medium. After 72 hours of culture, supernatants were collected and analyzed for cytokines using mouse IL-17 ELISA kit (Abcam, UK). Also, for investigation of the level of overexpression, supernatants from transduced rWJSCs were evaluated using enzyme-linked immunosorbent assay (ELISA). After 72 hours of culturing of transduced rWJSCs, supernatants were collected and analyzed for cytokines production using IL-4, IL-10 and LIF ELISA Kits (Abcam, UK). Fifty five days post-immunization, blood samples were collected from the mice to analyze circulating levels of IL-4, IL-10, LIF and IL-17 by ELISA assay according to the manufacturer’s protocol (Abcam, UK). The concentration of each cytokine was calculated based on the plotted standard curve.

MCF-7 cells were purchased from the Pasture Institute of Iran (Tehran, Iran). Dulbecco’s modified Eagle’s medium (DMEM), Fetal Bovine Serum (FBS), penicillin, and streptomycin were purchased from Life Technologies Co. (Gibco Life Technologies, Carlsbad, CA). Ketamine and xylazine were purchased from Sigma-Aldrich Co. (St Louis, MO, USA). The Cobalt-60 facility was provided by the Cancer Institute of Imam Khomeini hospital Complex of Iran (Tehran University of Medical Sciences, Tehran, Iran). Mouse monoclonal antibodies of anti-CD19, anti-CD4, and anti-CD8 were purchased from Abcam CO. INF-γ Mouse ELISA Kit (Abcam; ab46081), IL-4 Mouse ELISA Kit (Abcam; ab100710), and IL-17 Mouse ELISA Kit (Abcam; ab100702) were used for IFN-γ, IL-4, and IL-17 serum level assessment. Total RNA was isolated with a Total RNA extraction kit (GRM1002, VIOGENE, Sunnyvale, CA, USA). Template cDNAs were synthesized using SuperScript III (18080-051, Invitrogen, Carlsbad, CA). Quantitative RT-PCR was performed with a quantitative PCR mix (QPK-201T, Toyobo, Osaka, Honshu, Japan) using a real-time PCR detection system (CFX96; Bio-Rad Laboratories).