Overview

  • Product name
    Glycogen Assay Kit II (Colorimetric)
    See all Glycogen kits
  • Sample type
    Tissue, Adherent cells, Suspension cells
  • Assay type
    Quantitative
  • Sensitivity
    > 4 µg/ml
  • Range
    4 µg/ml - 40 µg/ml
  • Species reactivity
    Reacts with: Other species, Mammals
  • Product overview

    Glycogen Assay Kit II (Colorimetric) (ab169558) provides a simple, fast and robust way to measure Glycogen levels in various biological samples. This assay is suitable for measuring Glycogen levels in samples that contain reducing substances, which may interfere with the oxidase-based assays. In this assay, Glycogen is hydrolyzed into glucose, which is oxidized to form an intermediate that reduces a colorless probe to a colored product with strong absorbance at 450 nm. This high-throughput suitable assay kit can detect 4 - 40 µg/ml of Glycogen in samples.


    Review our Metabolism Assay Guide to learn about assays for metabolites, metabolic enzymes, mitochondrial function, and oxidative stress, and also about how to assay metabolic function in live cells using your plate reader.

  • Notes

    Glycogen serves as the main carbohydrate storage in animals and can be converted to glucose readily. It is primarily found in the liver and muscle tissues. Glycogen is a branched biopolymer comprising of α-1,4 linkage with α-1,6 linkages occurring every 8-10 glucose units along the backbone. Abnormal ability to utilize glycogen is found in diabetes and in several genetic glycogen storage diseases.

Properties

  • Storage instructions
    Store at -20°C. Please refer to protocols.
  • Components Identifier 100 tests
    Development Enzyme Mix (lyophilized) Green 1 vial
    Glycogen Development Buffer 1 x 25ml
    Glycogen Hydrolysis Buffer 1 x 25ml
    Glycogen Standard (2.0 mg/ml) Yellow 1 x 100µl
    Hydrolysis Enzyme Mix (lyophilized) Blue 1 vial
    Probe 1 vial
  • Research areas
  • Relevance
    Glycogen is the primary short term energy storage molecule in animals. It is synthesized primarily in the liver and muscle. Glycogen is a highly branched polymer of glucose molecules, connected with an alpha-1,4 linkage, branching via an alpha-1,6 linkage. Abnormal ability to utilize glycogen is found in diabetes and in several genetic glycogen storage diseases.

Images

  • Glycogen measured in cell lysates showing quantity (�g) per 1 mln cells.

    Samples with the concentration of 1e7 cells/mL (HeLa) and 6.6e6 cells/mL (HCT116) were used. Samples were diluted 2-6 fold.

  • Glycogen measured in tissue lysates showing quantity (�g) per mg of extracted protein.

    Protein concentration for samples varied from 25 mg/mL to 50 mg/mL. Samples were diluted 2-54 fold.

  • Standard curve: mean of duplicates (+/- SD) with background reads subtracted

Protocols

References

This product has been referenced in:
  • Granatiero V  et al. Role of p66shc in skeletal muscle function. Sci Rep 7:6283 (2017). Functional Studies ; Mouse . Read more (PubMed: 28740219) »
  • Iuchi H  et al. Time-dependent effects of ipragliflozin on behaviour and energy homeostasis in normal and type 2 diabetic rats: continuous glucose telemetry analysis. Sci Rep 7:11906 (2017). Functional Studies ; Rat . Read more (PubMed: 28928461) »

See all 4 Publications for this product

Customer reviews and Q&As

Abreviews
1) Dechorionate the embryos mechanically at 24 hpf
2) Remove the yolk
• Protocol for deyolking:
- Put the 10 dechorionated embryos in an eppendorf tube (1.5 ml) using a plastic
pipette, and place on ice to sedate the embryos.
- Take the eppendorf tube containing the embryos from ice, remove as much fluid (fish water/egg water) as possible and add deyolking buffer (*).
- Remove the yolks by triturating with a 200 µl pipette tip.
- Let the embryos float down to the bottom of the tube, and remove as much
deyolking fluid as possible.
- Rince twice with ice cold Ringer’s solution.
- Remove as much liquid as possible and put closed eppendorf tube in liquid nitrogen for 10 min, and store at -80ºC.
Or: you can proceed immediately with the protocol for Glycogen Assay Kit II analysis.
1. Glycogen Standard Curve:
Dilute Glycogen Standard to 0.2 mg/ml (0.2 µg/μl) by adding 10 µl of 2 mg/ml Glycogen Standard to 90 µl dH2O, mix well. Add 0, 2, 4, 6, 8 and 10 µl of 0.2 mg/ml Glycogen Standard into series of wells in 96 well plate to generate 0, 0.4, 0.8, 1.2, 1.6
and 2 µg/well Glycogen Standard. Adjust volume to 50 µl per well with Glycogen Hydrolysis Buffer.
2. Sample preparation:
10 zebrafish embryos should be rapidly homogenized with 200 µl ddH2O for 10 minutes on ice. Boil the homogenates for 10 minutes to inactivate enzymes. Centrifuge at 14500 rpm for 15 minutes and remove insoluble material. Collect the supernatant. Supernatant is ready to be assayed. Add 10-50 µl samples (~50 µg) into a 96 well plate and bring the volume to 50 µl with Glycogen Hydrolysis Buffer.
3. Hydrolysis:
Add 2 µl of Hydrolysis Enzyme Mix to Standard and samples, mix well. Incubate at room temperature for 30 minutes.
Note: Don’t add Hydrolysis Enzyme Mix to the sample background control.
4. Reaction Mix:
Mix enough reagents for the number of samples and standards to be performed. For each well, prepare a total 50 µl Reaction Mix containing:

Add 48 µl of the Reaction Mix to each well containing the Standard and samples and 50 µl of Background Control Mix to background control well.
5. Measurement:
Incubate at room temperature for 30 minutes. Measure OD450 nm with a microplate reader.
6. Data Analysis
Calculation: Subtract 0 Glycogen Standard reading from all readings. Plot the Glycogen Standard curve. If background control reading is significant, subtract the background control reading from sample reading. Apply the corrected sample reading to the Glycogen Standard curve to get B µg of Glycogen in the sample.

Figure 1 (A) Column 1: ug per well Glycogen Standard, Column 2 and 4: absorbance (Abs) values from first and second lecture, Column 3 and 5: absorbance less background Column 6: media of absorbance values. (B) Glycogen Standard Curve.

7. Results
To achieve the quantity of glycogen (ug) present in our sample, we applied the formula obtained from the standard curve plot:
y = 0,0876x – 0,0828
x = (y + 0,0828) / 0,0876 = glycogen amount from Standard Curve (B)
Sample Glycogen Concentration (C) = B (ug) / Sample volume used in the reaction (ul)

Figure 2 (A) Column 2 and 4: absorbance (Abs) values from first and second lecture, Column 3 and 5: absorbance less background Column 6: glycogen values obtained applying the formula: C = B (ug) / Sample volume used in the reaction (ul).
(B) Glycogen present in zebrafish embryos at 24 hours post fertilization (hpf).

The embryos used were zebrafish from AB strain line, breeded in fish water.

(*): DEYOLKING BUFFER
To obtain 400ml di ½ Ginzburg fish Ringer solution (deyolking buffer) weigh: 1,3 g NaCl; 0,05 g KCl; 0,06 g CaCl2 or 0,08 CaCl2 2H2O; Add milliQ water almost till 400 ml; Add 0,04 g of NaHCO3; Bring to volume (400 ml) with milliQ water; Filter and store at RT.
Username

Dr. Cinzia Bragato

Verified customer

Submitted Jan 17 2018

Thank you for your enquiry.

I can confirm that the kit is not species specific so it should be suitable for mouse. It has also been tested for tissue samples, so this wil also be covered by the guarantee.


https://www.abcam.com/co...

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