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Amplite® Colorimetric Glycogen Assay Kit *Red Color*
Product key features
The Amplite® Colorimetric Glycogen Assay Kit offers a fast, convenient, and sensitive method for quantifying glycogen in biological samples.
- Simple and direct detection: Employs an enzymatic hydrolysis step followed by a colorimetric reaction to specifically measure glycogen content in samples.
- Red colorimetric readout: Produces a stable red-colored product with absorbance measurable at 575 nm.
- Broad sample compatibility: Compatible across various sample types, including tissues and cultured cells.
Product description
The Amplite® Colorimetric Glycogen Assay Kit provides a streamlined assay for quantifying glycogen using a sensitive and reliable two-step enzymatic process. In the first step, glycogen is enzymatically hydrolyzed into glucose. In the second step, the released glucose undergoes an oxidation reaction, producing hydrogen peroxide, which then reacts with the proprietary Amplite® Red dye to generate a red-colored product. The intensity of this color is directly proportional to the glycogen concentration and can be easily measured at 575 nm using a standard absorbance microplate reader.
This kit is compatible with a wide range of biological matrices, including cell lysates, tissue extracts, and serum samples. The protocol is easy to follow, and it produces a stable colorimetric signal, enabling flexible endpoint detection for high-throughput screening or routine glycogen analysis in metabolic research, diabetes studies, and other related applications.
Example protocol
AT A GLANCE
- Prepare test samples along with serially diluted glycogen standards (40 μL).
- Add glycogen hydrolysis enzyme working solution (10 μL).
- Incubate at RT for 30 minutes.
- Add glycogen assay working solution (50 μL).
- Incubate at RT for 30 minutes.
- Measure the absorbance at 575 nm.
Note: Thaw all the kit components at room temperature before starting the experiment.
PREPARATION OF STOCK SOLUTIONS
Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles
Add 100 µL of DMSO (Component G) into Amplite® Red (Component A) to make 100X Amplite™ Red stock solution
Add 100 µL distilled water to Glycogen Hydrolysis Enzyme (Component C) to make a 20X Glycogen Hydrolysis Enzyme stock solution.
Add 100 µL distilled water to glycogen assay enzyme (Component E) to make 50X Glycogen Assay Enzyme Stock Solution.
PREPARATION OF STANDARD SOLUTIONS
https://www.aatbio.com/tools/serial-dilution/40013
PREPARATION OF WORKING SOLUTION
Add 10 µL of 20X Glycogen Hydrolysis Enzyme stock solution to 190 µL of Glycogen Hydrolysis Buffer (Component B).
Note: 200 µL Glycogen Hydrolysis Enzyme working solution is for 20 assays with 96-well plates. Please prepare the volume as needed proportionally. The working solution is not stable, prepare it freshly, use promptly and avoid direct exposure to light.
Add 10 µL of 100X Amplite® Red stock solution and 20 µL of 50X Glycogen Assay Enzyme Stock Solution to 970 µL of Glycogen Assay Buffer (Component D).
Note: 1mL Glycogen Assay Enzyme Working Solution is for 20 assays with 96-well plates. Please prepare the volume as needed proportionally. The working solution is not stable, use it promptly and avoid direct exposure to light.
SAMPLE EXPERIMENTAL PROTOCOL
- Prepare Glycogen Standards (STD1-7), blank controls (BL), and test samples (TS) according to the layout provided in Tables 1 and 2. For a 96-well plate, add 40 µL per well, and for a 384-well plate, use 20 µL per well instead of 40 µL.
- Add 10 µL of Glycogen Hydrolysis Enzyme working solution to each well of blank control, glycogen positive control and test samples. For a 384-well plate, add 5 µL working solution into each well instead.
- Incubate at RT for 30 minutes.
- Add 50 µL of Glycogen Assay Enzyme working solution to each well of blank control, glycogen positive control and test samples. For a 384-well plate, add 25 uL working solution into each well instead.
- Incubate at RT for 30 minutes.
- Measure absorbance at 575 nm.
Table 1. Layout of glycogen standards and test samples in a clear bottom 96-well microplate. STD = glycogen standards (STD1-STD7, 0.6 to 40 µg/ml), BL = Blank Control, TS = Test Samples.
BL | BL | Positive Control | Cell content |
STD 1 | STD 1 | ... | ... |
STD 2 | STD 2 | ... | ... |
STD 3 | STD 3 | ||
STD 4 | STD 4 | ||
STD 5 | STD 5 | ||
STD 6 | STD 6 | ||
STD 7 | STD 7 |
Table 2. Reagent composition for each well.
Well | Volume | Reagent |
STD1-STD7 | 40 µL | Serial Dilutions (0.6 to 40 µg/ml) |
BL | 40 µL | Hydrolysis buffer |
TS | 40 µL | Test Sample |
References
Authors: Llauradó, A and Pinós, T and Codina-Solà, M and Martínez-Saez, E and Restrepo-Vera, J L and Salvadó, M and Sanchez-Tejerina, D and Sotoca, J and Muñoz, P and Rovira-Moreno, E and Büyükdereli, Leyla and Garcia-Arumi, E and Vidal-Taboada, J M and Ovelleiro, D and Juntas-Morales, R
Journal: Molecular genetics and metabolism (2025): 109140
Authors: Fu, Jingjing and Wang, Weijun and Sun, Youmei and Zhang, Yousen and Luo, Qihao and Wang, Zhongping and Wang, Degang and Feng, Yanwei and Xu, Xiaohui and Cui, Cuiju and Sun, Guohua and Li, Zan and Yang, Jianmin
Journal: Foods (Basel, Switzerland) (2024)
Authors: Chen, Siyu and Bouchibti, Yasmine and Xie, Yixuan and Chen, Ye and Chang, Vincent and Lebrilla, Carlito B
Journal: Analytical chemistry (2023): 12884-12892
Authors: Li, Meiqin and Li, Zhoulei and Wei, Luyong and Li, Lujie and Wang, Meng and He, Shaofu and Peng, Zhenpeng and Feng, Shi-Ting
Journal: Quantitative imaging in medicine and surgery (2023): 4933-4942
Authors: Young, Lyndsay E A and Conroy, Lindsey R and Clarke, Harrison A and Hawkinson, Tara R and Bolton, Kayli E and Sanders, William C and Chang, Josephine E and Webb, Madison B and Alilain, Warren J and Vander Kooi, Craig W and Drake, Richard R and Andres, Douglas A and Badgett, Tom C and Wagner, Lars M and Allison, Derek B and Sun, Ramon C and Gentry, Matthew S
Journal: EMBO molecular medicine (2022): e16029
Product protocol