Ultrasensitive mouse insulin enzyme linked immunosorbent assay elisa kit

The mouse GFP⁺-iPSC-derived β-like cells were characterized by immunofluorescence and glucose-stimulated insulin secretion. For immunofluorescence, cells were fixed by 4% paraformaldehyde (PFA) (Solarbio, Beijing, China) for 20 min, permeabilized by 0.1% Triton X-100 (Solarbio) for 10 min, and blocked with 5% bovine serum albumin (BSA) (Solarbio) for 30 min. After that, the cells were incubated with rabbit anti-insulin 1:100 (Abcam, Cambridge, MA, USA) or rabbit anti-C-peptide 1:100 (Abcam) overnight at 4 °C. The next day, the cells were washed 3 times with phosphate-buffered saline (PBS) and incubated with Alexa Flour 594-conjugated goat anti-rabbit secondary antibody 1:500 (Abcam) at room temperature for 1 h. Subsequently, the cells were stained with Hoechst (Sigma-Aldrich) for 15 min and washed with PBS 3 times. The cells were visualized by an Olympus fluorescence microscope (Olympus, Tokyo, Japan).
For a glucose-stimulated insulin secretion assay, the iPSC-derived β-like cells were exposed to a glucose gradient (0, 5, 15, 30, and 45 mM), and insulin secretion was measured by an ultrasensitive mouse insulin enzyme-linked immunosorbent assay (ELISA) kit (Mercodia, Uppsala, Sweden).

Insulin levels and adiponectin levels were measured by Ultra-Sensitive Mouse Insulin Enzyme-Linked Immunosorbent Assay (ELISA) kit (Mercodia) and adiponectin kits (Abcam) according to the manufacturer’s instruction. The plates were read with a microplate reader (SpectraMax MS; Molecular Devices). The coefficient of variance between replicates was 4.5% and 4.3%, and the corresponding detection range was 1.56 to 100 ng/mL and 62.5 to 4000 pg/mL. Blood samples were collected from tail. For measurement of insulin and adiponectin levels in SM, tissues were first lysed with RIPA lysis buffer (Biovision) and centrifuged at 12 000g at 4 °C for 20 minutes. The supernatants were collected for analysis.