Cell mitochondrial stress test kit

By cooperating with KWINBON (Beijing, China), mouse monoclonal anti-MERS-CoV nsp1 antibody was prepared in our laboratory; Rabbit monoclonal anti-LAMP1 antibody (6502, CST); Rabbit polyclonal anti-RPS18 antibody (A11687, Abclonal); Rabbit polyclonal anti-LSM14A antibody (A16682, Abclonal); Rabbit polyclonal anti-G3BP1 antibody (A14836, Abclonal); Rabbit polyclonal anti-COX4I1 antibody (A6564, Abclonal); TRIzol™ Reagent (15,596–026, Invitrogen); RNase inhibitor (R0102, Beyotime); Protease inhibitor (B14001, bimake); Phosphatase inhibitor (B15001, bimake); DNase I (D7073, Beyotime); Proteinase K (TIANGEN, RT403); poly(I:C) (APExBIO, B5551); D-galactosamine (Beyotime, ST1213); Tetracyclin (T8180, Solarbio); Mito-Tracker Red CMXRos (Beyotime, C1071S); Cell Counting Kit-8 (CCK-8) (K1018, APExBIO); Cell Cycle Analysis Kit (CY2001, Tianjin Sungene Biotech Co, Ltd); Cell Mitochondrial Stress Test Kit (103,015–100, Agilent Seahorse); Mitochondrial membrane potential assay kit with JC-1 (C2006, Beyotime).

A Seahorse Mito Fuel Flex Test kit and cell mitochondrial stress test kit were used to measure the oxygen consumption rates (OCR) in CD3⁺ T cells according to the manufacturer’s protocol (Agilent, Santa Clara, CA). Purified T cells were seeded in each well of an XFp cell culture mini plate at a density of 2 × 10⁵ cells/well. The XFp cell culture plates were pre-coated with CellTak (Corning, NY, USA) to allow the T cells to adhere to the bottom of wells. Measurements were performed on a Seahorse XFp Analyzer (Agilent, Santa Clara, CA, USA). The Seahorse Wave software was used to interpret the acquired data and calculate the OCR of the purified T cells.