Lipids were extracted from liver tissue using chloroform:methanol mixture (2:1 v/v) (40 (link)). enzymatic assay kits were used to measure hepatic and plasma concentrations of triglyceride (Wako Diagnostics). Plasma AST and ALT activities were measured by an enzymatic assay kit (Thermo Fisher Scientific). Plasma insulin and Fgf21 concentrations were determined using commercially available mouse ELISA kits from Crystal Chem and R&D Systems, respectively, according to the manufacturer's specification. Protein concentrations were determined using a bicinchoninic acid assay (Thermo Fisher Scientific).
Mouse elisa kit
Manufactured by Crystal Chem
Sourced in United States
The Mouse ELISA Kit is a laboratory equipment used for the detection and quantification of mouse-specific proteins or analytes in biological samples. It utilizes the Enzyme-Linked Immunosorbent Assay (ELISA) technique to measure the target analyte's concentration. The kit includes the necessary reagents, plates, and instructions to perform the assay.
Automatically generated - may contain errors
Lab products found in correlation
Colorimetric assay, by EKF Diagnostics (2 mentions)
Bicinchoninic acid assay, by Thermo Fisher Scientific (2 mentions)
Enzymatic assay kit, by Fujifilm (1 mentions)
Enzymatic assay kit, by Thermo Fisher Scientific (1 mentions)
Qiazol lysis reagent, by Qiagen (1 mentions)
Ab37593, by Abcam (1 mentions)
Nupage bis tris gel, by Thermo Fisher Scientific (1 mentions)
Irdye 800 coupled goat anti rabbit secondary antibody, by Rockland Immunochemicals (1 mentions)
Odyssey infrared imaging system, by LI COR (1 mentions)
Benchmark plus microplate reader, by Bio-Rad (1 mentions)
Humulin, by Eli Lilly (1 mentions)
Insulin, by Crystal Chem (1 mentions)
Glucometer, by Roche (1 mentions)
18 protocols using mouse elisa kit
1
Liver Lipid Profiling Protocol
2
Metabolic Profiling of Offspring
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Offspring were anesthetized with ketamine/xylazine (50 mg/mL; injected 0.1 cc per 10 g body weight [bw]), and fat mass and lean mass were measured in offspring by DEXA (Lunar PIXImus2 mouse densitometer). Body weight was measured every 2–3 days and reported every 12 weeks, and food consumption was measured weekly. Retro-orbital sinus bleeds were performed after an overnight fast (2200 h to 900 h); plasma insulin and leptin were measured using mouse ELISA kits (Crystal Chem Inc.); and triglyceride, cholesterol, and free fatty acid were measured by colorimetric assay (Stanbio). Muscle glycogen and triglycerides were determined in tibialis anterior muscles as previously described (23 (link)). Tibialis anterior muscle was used for RNA extraction using QIAzol Lysis Reagent (Qiagen). mRNA expression was measured by quantitative RT-PCR (primers in Supplementary Table 1 ). Tissue processing and immunoblotting were performed as previously described (24 (link)). Antibodies used were GLUT4 (AB1346) (Millipore) and HKII (AB37593) (Abcam).
3
Metabolic Profiling of Mouse Plasma and Liver
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Blood was collected from the retro-orbital sinus after an overnight fast (2200 to 0900 h). Plasma insulin and leptin were measured using mouse ELISA kits (Crystal Chem Inc., and triglyceride, cholesterol, and free fatty acid were measured by colorimetric assay (Stanbio Laboratory). mRNA levels of various liver genes were measured by quantitative RT-PCR using primers shown in Supplementary Table 1 . Liver triglycerides were measured as previously described (33 (link)). Tissue processing and immunoblotting were performed as previously described (34 (link)). Antibodies used were GLUT4 (AB1346; Millipore) and HKII (AB37593; Abcam).
4
Fasting-Induced Changes in Metabolic Markers
5
Serum Adipokine Quantification in Mice
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Blood insulin, leptin, and adiponectin concentrations were measured using an ELISA reader with corresponding mouse ELISA kits (CrystalChem, Inc., Elk Grove Village, IL, USA). Blood was collected from the inferior vena cava with a sterile syringe after the abdomen had been opened to analyze blood indices. The collected blood was allowed to stand for 1 h on ice and centrifuged at 3000 rpm (4 °C) for 10–15 min. The serum supernatant was diluted 10 times and a 50 μL aliquot was reacted at 4 °C for 12 h before incubation with HRP-anti-mouse insulin, leptin, and adiponectin antibodies at 4 °C for 4 h. Non-specifically bound antibodies were removed by washing with phosphate buffered saline-tween (PBS/T) twice. After treatment with substrate solution for 30 min, absorbance was measured at 450 and 630 nm using the Benchmark Plus microplate reader (BioRad, Hercules, CA, USA).
6
Plasma Analysis of Metabolic Markers
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Plasma samples were prepared at the time of sacrifice as previously described (Haviland et al. 2012 ). Plasma insulin and leptin levels were measured using mouse ELISA kits (Crystal Chem, Downers Grove, IL). Adiponektin level was measured by adiponectin EIA kit (ACRP30, Norcross, GA).
7
Metabolic Profiling of Offspring
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Metabolic testing was performed on male and female offspring at various time points throughout 52 weeks of life. Intraperitoneal glucose tolerance tests (IPGTTs) and insulin tolerance tests (ITTs) were performed as previously described (30 (link),31 (link)). Blood was collected from the retro-orbital sinus after an overnight fast (2200–0900 h) to measure plasma insulin levels using a mouse ELISA kit (Crystal Chem Inc). The assessment of fat and lean mass was performed using DEXA (Lunar PIXImus2 mouse densitometer) with offspring anesthetized with an injection of a mixture of ketamine HCL (100 mg/kg i.p.) and xylazine (10 mg/kg i.p.) before scanning.
8
Measuring Leptin Levels in Mice
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Plasma levels of leptin were determined by a mouse ELISA Kit (Crystal Chem, Zaandam, The Netherlands), following the manufacturer’s instructions.
9
Glucose and Insulin Tolerance Tests in Mice
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For GTT, mice were fasted for 16 h (1700 to 0900 h) with free access to drinking water. A baseline blood sample was collected from the tails of fully conscious mice, followed by an intraperitoneal injection of glucose (0.75 g glucose/kg body weight), and blood was taken from the tails for glucose measurements at 0, 15, 30, 60, and 120 min. For insulin tolerance tests, mice were fasted for 6 h (0900 to 1500 h), and baseline blood samples were collected from the tails of fully conscious mice. insulin (1 unit/kg body weight) (Humulin; Eli Lilly and Company, Indianapolis, IN, US) was administered by intraperitoneal injection, and blood samples were taken from the tail at 0, 30, 60, 90 and 120 min postinjection. Blood was collected from the retroorbital sinus after an overnight fast (16 h) to measure plasma insulin levels using a mouse ELISA kit (Crystal Chem Inc.).
10
Plasma Leptin Quantification by ELISA
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Plasma was obtained after blood centrifugation (2000 g, 15 min). Plasma levels of leptin were determined by mouse ELISA kit (Crystal Chem, Zaandam, Netherlands), following the manufacturer's instructions.
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