Glycogen Assay Kit

Catalog No. ABIN1000282

Product Details

Target: Glycogen

Application: Biochemical Assay (BCA)

Specificity: 2 μg/mL

Characteristics: Use as little as 10 µL samples. Linear detection range: 2 to 200 µg/mL glycogen for colorimetric assays and 0.2 to 20 µg/mL for fluorimetric assays.

Components: Assay Buffer: 12 mL. Enzyme A: 120 µL. Enzyme B: 120 µL. Dye Reagent: 120 µL. Standard: 50 µL 50 mg/mL.

Material not included: Pipeting devices, centrifuge tubes, clear flat bottom 96-well plates and plate reader.

Application Details

Comment:

1. If the sample contains glucose, a Sample Blank well should be added: Prepare Sample Blank reagent by mixing 90 μL Assay Buffer, 1 μL Enzyme B, and 1 μL Dye Reagent (No Enzyme A). Add this reagent only to the Sample Blank wells. Subtract the OD or fluorescence of the Sample Blank from the sample readings to calculate glycogen concentration. 2. This assay is based on a kinetic reaction, the use of a multi- channel pipettor for adding the working reagent is recommended. 3. Interference. SH-group containing reagents (e.g., DTT, - mercaptoethanol) may interfere with this assay and should be avoided in sample preparation.

Protocol: 1. Equilibrate all components to room temperature. During experiment, keep thawed enzymes in a refrigerator or on ice.
2. Standards and samples: Dilute standard by mixing 5 µL Standard with 1.245 mL dH2O to give 200 µg/mL standard. Dilute standard in dH2O. Transfer 10 µL standard and samples into separate wells of a clear flat-bottom microplate.
3. Working Reagent. For each reaction well, mix 90 µL Assay Buffer, 1 µL Enzyme A, 1 µL Enzyme B and 1 µL Dye Reagent in a clean tube. Transfer 90 µL Working Reagent into each reaction well. Tap plate to mix.
4. Incubate 30 min at room temperature. Read optical density at 570nm (550-585nm).

Calculation of Results:

Subtract Blank reading (OD570nm or fluorescence intensity) from the standard reading values and plot the OD or F against standard concentrations.

Restrictions: For Research Use only

Handling

Storage: -20 °C

References

Tucci, Pearson, Herebian, Spiekerkoetter: "Long-term dietary effects on substrate selection and muscle fiber type in very-long-chain acyl-CoA dehydrogenase deficient (VLCAD(-/-)) mice." in: Biochimica et biophysica acta, Vol. 1832, Issue 4, pp. 509-16, (2013) (PubMed).

Anderson, Politch, Pudney, Marquez, Snead, Mauck: "A quantitative glycogen assay to verify use of self-administered vaginal swabs." in: Sexually transmitted diseases, Vol. 39, Issue 12, pp. 949-53, (2012) (PubMed).

Yin, van der Horst, van Melle, Qian, van Gilst, Silljé, de Boer: "Metformin improves cardiac function in a nondiabetic rat model of post-MI heart failure." in: American journal of physiology. Heart and circulatory physiology, Vol. 301, Issue 2, pp. H459-68, (2011) (PubMed).

Iwabuchi, Kawahara: "Inducible astrocytic glucose transporter-3 contributes to the enhanced storage of intracellular glycogen during reperfusion after ischemia." in: Neurochemistry international, Vol. 59, Issue 2, pp. 319-25, (2011) (PubMed).

Bouitbir, Charles, Rasseneur, Dufour, Piquard, Geny, Zoll: "Atorvastatin treatment reduces exercise capacities in rats: involvement of mitochondrial impairments and oxidative stress." in: Journal of applied physiology (Bethesda, Md. : 1985), Vol. 111, Issue 5, pp. 1477-83, (2011) (PubMed).

Primassin, Tucci, Spiekerkoetter: "Hepatic and muscular effects of different dietary fat content in VLCAD deficient mice." in: Molecular genetics and metabolism, Vol. 104, Issue 4, pp. 546-51, (2011) (PubMed).

Target Details

Target: Glycogen

Abstract: Glycogen Products

Background: Quantitative determination of glycogen by colorimetric (570nm) or fluorometric (585/530nm) methods.
Procedure: 30 min.

Glycogen is a branched polysaccharide of glucose units linked by alpha-1,4 glycosidic bonds and alpha-1,6 glycosidic bonds. It is stored primarily in the liver and muscle, and forms an energy reserve that can be quickly mobilized to meet a sudden need for glucose. The most common glycogen metabolism disorder is found in diabetes, in which, due to abnormal amounts of insulin, liver glycogen can be abnormally accumulated or depleted. Genetic glycogen storage diseases have been associated with various inborn errors of metabolism caused by deficiencies of enzymes necessary for glycogen synthesis or breakdown. Simple, direct and automation-ready procedures for measuring glycogen concentrations find wide applications in research and drug discovery. This glycogen assay uses a single Working Reagent that combines the enzymatic break down of glycogen and the detection of glucose in one step. The color intensity of the reaction product at 570nm or fluorescence intensity at gamma em/ex = 585/530nm is directly proportional to the glycogen concentration in the sample. This simple convenient assay is carried out at room temperature and takes only 30 min.