Mouse il 1β il 1f2 quantikine elisa kit

Mouse BMDMs and hMDMs were infected with C. albicans (MOI = 10) for the indicated time periods. IL-1β in culture supernatants was measured with a mouse IL-1β/IL-1F2 Quantikine ELISA Kit or human IL-1 beta/IL-1F2 DuoSet ELISA Kit (R&D Systems). To measure intracellular IL-1β, BMDMs were collected and lysed with PBS/0.5% Triton X-100. Proteins in the supernatants were precipitated with chloroform/methanol (1:4). Unattached cells (<5%) were removed by washing twice with PBS. The levels of IL-1β in each sample were measured using the ELISA kit as described above.

The levels of myeloperoxidase (MPO) and interleukin (IL)-1β in the colonic tissues were measured using the mouse MPO enzyme-linked immunosorbent assay (ELISA) kit (HK210, Hycult Biotechnology) and mouse IL-1β/IL-1F2 Quantikine ELISA kit (R&D Systems Inc.), respectively. All assays were performed in triplicates.

In order to validate the IL-1αβ gene deficiency in mice, plasma and brain IL-1β levels were examined under basal conditions and after endotoxin challenge. WT and IL-1αβKO mice were treated i.p. with 20 mg kg⁻¹ LPS. Six hours after LPS injection, mice were euthanized by an anesthetic overdose of the isoflurane inhalation and approx. 800 μL blood and brain structures were collected. The brain was not perfused before tissue harvesting. Blood was taken from the vena cava with heparinized syringes inserting a 25 G × 5/8″ needle. PFC, HP and STR were homogenized in phosphate buffered saline. The concentration of IL-1β in the brain structures was determined using a specific commercially available Mouse IL-1β/IL-1F2 Quantikine ELISA Kit (R&D Systems, Minneapolis, MN, US, Cat# MLB00C) according to the manufacturer’s instructions. Samples were not diluted. Absorbance was measured at 450 nm. IL-1β concentration was expressed as pg mg¹ (link) protein. IL-1β levels were normalized to total protein levels measured photometrically using the BCA Protein Assay Kit (Thermo Scientific Pierce, Rockford, IL, US, Cat# 23227). Absorbance was measured at 560 nm using a Cytation 5 Cell Imaging Multi-Mode Reader (BioTek, Winooski, VT, US).

Blood from C5BL/6 mice with tumor size of 20–40 mm³ was harvested at the selected time points following single intratumoral injection of 300 μg of RT53 or vehicle (normal saline). In vivo secretion of IL-1β and IL-6 in the serum was assessed using the Mouse IL-1β/IL-1F2 Quantikine ELISA Kit and Mouse IL-6 Quantikine ELISA Kit, respectively, per the manufacturer's instruction (R&D System).

Mouse cytokine secretions were obtained from PBMC supernatant, including IL-1β (Interleukin 1β), IL-6, and monocyte chemotactic protein-1 (MCP1), and were measured using Mouse IL-1β/IL-1F2 Quantikine ELISA Kit (#MLB00C), Mouse IL-6 Quantikine ELISA Kit (#M6000B), and Mouse CCL2/JE/MCP1 Quantikine ELISA Kit (#MJE00B), respectively, according to manufacturers’ instructions from R&D Systems (27 (link)).