Glucagon ELISA Assay Kit

$650.00

The Eagle Biosciences Glucagon ELISA Assay kit is intended for the quantification of Glucagon in serum, plasma or cell culture supernatants. The Eagle Biosciences Glucagon ELISA Assay is for Research Use Only and should not be used for diagnostic procedures.

SKU: ARG81293 Categories: , ,

Glucagon ELISA Assay Kit

For Research Use Only

Size: 1×96 wells
Sensitivity: 15 pg/mL
Dynamic Range: 31.25-2000 pg/ml
Incubation Time: 5 hours
Sample Type: Serum, Plasma, Cell culture Supernatants
Sample Size: 100 µL

Additional Information

Assay Background

The protein encoded by this gene is actually a preproprotein that is cleaved into four distinct mature peptides. One of these, glucagon, is a pancreatic hormone that counteracts the glucose-lowering action of insulin by stimulating glycogenolysis and gluconeogenesis. Glucagon is a ligand for a specific G-protein linked receptor whose signalling pathway controls cell proliferation. Two of the other peptides are secreted from gut endocrine cells and promote nutrient absorption through distinct mechanisms. Finally, the fourth peptide is similar to glicentin, an active enteroglucagon. Glucagon plays a key role in glucose metabolism and homeostasis. Regulates blood glucose by increasing gluconeogenesis and decreasing glycolysis. A counter regulatory hormone of insulin, raises plasma glucose levels in response to insulin-induced hypoglycemia. Plays an important role in initiating and maintaining hyperglycemic conditions in diabetes. GLP-1 is a potent stimulator of glucose-dependent insulin release. Play important roles on gastric motility and the suppression of plasma glucagon levels. May be involved in the suppression of satiety and stimulation of glucose disposal in peripheral tissues, independent of the actions of insulin. Have growth-promoting activities on intestinal epithelium. May also regulate the hypothalamic pituitary axis (HPA) via effects on LH, TSH, CRH, oxytocin, and vasopressin secretion. Increases islet mass through stimulation of islet neogenesis and pancreatic beta cell proliferation. Inhibits beta cell apoptosis. GLP-2 stimulates intestinal growth and up-regulates villus height in the small intestine, concomitant with increased crypt cell proliferation and decreased enterocyte apoptosis. The gastrointestinal tract, from the stomach to the colon is the principal target for GLP-2 action. Plays a key role in nutrient homeostasis, enhancing nutrient assimilation through enhanced gastrointestinal function, as well as increasing nutrient disposal. Stimulates intestinal glucose transport and decreases mucosal permeability. Oxyntomodulin significantly reduces food intake. Inhibits gastric emptying in humans. Suppression of gastric emptying may lead to increased gastric distension, which may contribute to satiety by causing a sensation of fullness. Glicentin may modulate gastric acid secretion and the gastro-pyloro-duodenal activity. May play an important role in intestinal mucosal growth in the early period of life.

Assay Principle

This assay employs the quantitative sandwich enzyme immunoassay technique. An antibody specific for Glucagon has been pre-coated onto a microtiter plate. Standards or samples are pipetted into the wells and any Glucagon present is bound by the immobilized antibody. After washing away any unbound substances, a biotin-conjugated antibody specific for Glucagon is added to each well and incubate. Following a washing to remove unbound substances, streptavidin conjugated to Horseradish Peroxidase (HRP) is added to each microplate well and incubated. After washing away any unbound antibody-enzyme reagent, a substrate solution (TMB) is added to the wells and color develops in proportion to the amount of Glucagon bound in the initial step. The color development is stopped by the addition of acid and the intensity of the color is measured at a wavelength of 450nm ±2nm.The concentration of Glucagon in the sample is then determined by comparing the O.D of samples to the standard curve. 

Assay Procedure

1. Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal it.

2. Add 100 μl of standards, samples and zero controls (Standard/Sample diluent) into wells. Incubate on a microplate shaker at 700 rpm for 120 minutes at RT.

3. Aspirate each well and wash, repeating the process three times for a total four washes. Wash by filling each well with 1× Wash Buffer (350 μl) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each is essential to good performance. After the last wash, remove any remaining Wash Buffer by aspirating, decanting or blotting against clean paper towels.

4. Add 100 μl 1X Antibody conjugate into each well. Cover wells and incubate on a microplate shaker at 700 rpm for 120 minutes at RT.

5. Aspirate each well and wash as step 3.

6. Add 100 μl of 1X HRP-Streptavidin solution to each well. Cover wells and incubate on a microplate shaker at 700 rpm for 20 minutes at RT.

7. Aspirate each well and wash as step 3.

8. Add 100 μl of TMB Reagent to each well. Incubate for 20-25 minutes at RT in dark.

9. Add 50 μl of Stop Solution to each well. The color of the solution should change from blue to yellow. Gently tap the plate to ensure thorough mixing

10. Read the OD with a microplate reader at 450nm immediately. (Optional: read at 610-650 nm as the reference wave length). It is recommended read the absorbance within 5 minutes after adding the stop solution. 

Manual

Product Manual