1 2 3 ultrasensitive mouse insulin elisa kit

Glucose-stimulated insulin secretion assay was performed according to previously established methods (Pagliuca et al. 2014) (link). Briefly, PE clusters were washed with Krebs buffer and were then pre-incubated in 2 mM glucose for 2 h to remove residual insulin. Clusters were then washed twice, incubated in 2 mM glucose for 30 min, and supernatant collected. Then clusters were washed again, incubated in 20 mM glucose for another 30 min, and supernatant collected. Clusters were then dispersed into single cells and cell number was counted. Supernatant samples containing secreted insulin were processed using the 1-2-3 UltraSensitive Mouse Insulin ELISA Kit (ALPCO Diagnostics, Salem, NH, USA).

Body weight, blood glucose, and plasma insulin were measured prior to PE transplant (day 0), 4 weeks after transplant (day 28), gestation day 0 (GD0), GD10, and GD20. Body weight was measured on a top-loading balance (Accu-622; Fisher Scientific, Waltham, MA, USA), and nonfasting maternal blood samples were obtained via tail venipuncture to determine insulin and glucose levels. Blood glucose levels were determined by glucometer (Lifescan Surestep, Fisher Scientific). Plasma insulin levels were quantified by 1-2-3 UltraSensitive Mouse Insulin ELISA Kit (ALPCO Diagnostics), which detects both insulin 1 and 2.