Rat mouse proinsulin elisa kit

For pancreatic islet isolation, ice-cold 0.45 mg/mL collagenase type V buffer (Sigma-Aldrich, St. Louis, MO) (in Hanks’ balanced salt solution without Ca²⁺) was perfused into the bile duct. The pancreas was immediately dissected out and incubated at 37°C for 10–11 min in a water bath for digestion, and islets were washed twice in Hanks’ balanced salt solution and then handpicked under a stereo microscope. Islets were incubated overnight in RPMI complete media supplemented with 10% FCS, 100 units/mL penicillin, and 100 μg/mL streptomycin (Thermo Fisher Scientific). For GSIS, islets were preincubated in Krebs-Ringer buffer solution (bicarbonate HEPES) containing 2% BSA and 2.8 mmol/L glucose for 1 h. Islets were then separated into groups of 10 and incubated with either 2 or 20 mmol/L glucose for 1 h, and media were collected for GSIS. Total insulin content was extracted from 10 islets in 0.2 N/80% acid ethanol. Insulin released into the medium and total insulin content were measured using a rat/mouse insulin ELISA kit (Crystal Chem) (25 (link)). Total proinsulin content was measured using a rat/mouse proinsulin ELISA kit (Mercodia).

Intraperitonal glucose tolerance tests (IPGTTs), intraperitoneal insulin tolerance tests (IPIITTs), oral glucose tolerance tests (OGTTs), and plasma insulin measurements were performed as previously described (24 (link)). Plasma proinsulin levels were measured in fasted (16 h) animals using a rat/mouse proinsulin ELISA kit (Mercodia). Plasma β-ketones were measured from fed or fasted (16 h) mice using an Areo 2K device (GlucoMen, Berkshire, U.K.).