Raybio mouse insulin elisa kit

The serum glucose concentrations and the levels of total cholesterol and triglycerides were measured using the Fuji DRI-CHEM NX500V system (Fuji Film, Tokyo, Japan). The plasma insulin levels were determined using a RayBio mouse insulin ELISA kit (catalog number: ELM-Insulin, RayBiotech, Norcross, GA, USA). The plasma Angptl8 levels were measured using a mouse Betatrophin ELISA kit (catalog number: E11633m, Wuhan EIAab Science Co, Ltd., Wuhan, China). The absorbance measurement was performed using a Microplate Spectrophotometer (Epoch, BioTek, Winooski, VT, USA) at 450 nm wavelength.

The concentrations of insulin and glucagon were measured in the serum using ELISA kits (RayBio® Mouse Insulin ELISA Kit; RayBiotech Inc., Norcross, GA, USA, and Quantikine® ELISA Glucagon Immunoassay; R&D Systems) as previously described.⁶³ Serum samples were harvested at 6h and 7 days after CCI (n=3 per time point) in male and female.
White adipose tissue (WAT) was collected from the subcutaneous abdominal (inguinal) adipose panniculus. PPARγ and adiponectin levels, present in adipose tissue lysates, were measured using ELISA kit (MyBiosource Inc. San Diego USA and Quantikine® ELISA Immunoassay; R&D Systems respectively) in male and female mice (n=3 per time point), before and seven days post CCI. All Incubations and washes were performed following the manufacturer’s instructions. Then a substrate solution was added. The intensity of the colored product was directly proportional to the concentration of insulin and glucagon present in the sera and of PPARγ and adiponectin in adipose tissue and was read at 450 nm (with wavelength correction at 570 nm for insulin, glucagon and adiponectin detection).

All chemicals were obtained from Merck (Merck KGaA, Darmstadt, Germany). Monoclonal antibodies against pNF-kB p65 (27.Ser 536), NF-kB p65 (F-6), and PCNA (PC10) were purchased from Santa Cruz Biotechnology (Tebu Bio, Milan, Italy). Secondary anti-mouse conjugated to horseradish peroxidase (HRP) was obtained from Bio-Rad (Bio-Rad Laboratories, Inc., Hercules, CA, USA). The tert-butyl hydroperoxide (tBHP) was a kind gift from A. Signorile (University of Bari, Aldo Moro, Bari. Italy). The RayBio^® Mouse Insulin ELISA Kit was from RayBiotech (Norcross, GA, USA).