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| Sample Type(s) | Cells |
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Overview
For quantitative fluorescent immunoenzymatic assay of AMPK phosphorylation status in cultured cells. The assay uses Cell-Based ELISA (FL530/585 nm, 360/450 nm) for signal readout. Compatible sample input includes Cells. Typical stated assay timing is Assay takes 6.5 hrs, hands-on time 2.5 hrs.
Key elements and design rationale
- Readout format: Cell-Based ELISA (FL530/585 nm, 360/450 nm) supports plate-based signal acquisition and consistent comparison across matched samples.
- Sample compatibility: The stated sample scope includes Cells, which is useful when aligning matrix type with calibration and control design.
- Workflow timing: The listed assay time of Assay takes 6.5 hrs, hands-on time 2.5 hrs helps frame batch planning, replicate handling, and plate throughput.
- Feature emphasis: Sensitive. Can measure pAMPK modulation in as little as 500 cells/well.
Additional feature notes highlight New and improved. Total assay time reduced from the standard 21 hours to 6.5 hours (hands-on time 2.5 hrs); Simple and convenient. Cells are directly cultured in 96-well plates. No cell lysis is necessary. Available format information for this listing includes 100 Tests.
Biological background
This product is centered on measurement of enzyfluo ampk phosphorylation within the matrices described for the assay. In practice, datasets from this type of format are typically interpreted by comparing relative signal, activity, or abundance across matched control and experimental groups rather than relying on a single value in isolation. Careful alignment of sample matrix, incubation window, and calibration strategy is important when comparing results across plates, operators, or study days.
More details
The 5-AMP-activated protein kinase (AMPK) is a key sensor of intracellular energy balance. AMPK is activated in response to an increase in the AMP/ATP ratio which can be caused by a number of factors such as muscle contraction, starvation, or hypoxia. AMPK is a heterotrimeric protein complex comprising α- (63 kDa), β- (38 kDa) and γ- (38 kDa) subunits. For each subunit, isoforms have been identified (α1, α2, β- 1, β- 2, γ1, γ2, γ3) which theoretically allow the formation of 12 different proteins. The α-subunit contains a serine/threonine kinase domain and the regulatory subunits contain binding sites for AMP and ATP and for glycogen. AMPK is activated by phosphorylation on Thr-172 within the catalytic domain. AMP binding results in a 2 to 5-fold increase in AMPK activity compared to the basal level. The binding of AMP to the α-subunit causes allosteric activation of the kinase and induces a conformational change in the kinase domain that protects AMPK from dephosphorylation of Thr-172.BioAssay Systems cell-based ELISA measures phosphorylated AMPK in whole cells and normalizes the signal to the total protein content. The antibody recognizes both α-subunits and, thus, can be used for cells from all tissues (human, mouse, rat). This simple and efficient assay eliminates the need for cell lysate preparation and can be used to study AMPK regulation in short-term and long-term assays. In this assay, cells grown in 96-well plates are fixed and permeabilized in the wells. AMPK phosphorylation (pAMPK) is measured using a fluorescent ELISA followed by total protein measurement in each well.
Detection method
Cell-Based ELISA (FL530/585 nm, 360/450 nm).
Procedures and timing
Stated procedure or timing information: Assay takes 6.5 hrs, hands-on time 2.5 hrs.
Research relevance and current trends
- Plate-based quantification and side-by-side group comparison remain central use cases for this assay format.
- The product notes emphasize multi-sample throughput, making it relevant for screening-oriented and larger batch comparison studies.
- Short assay timing and plate compatibility support time-course or repeated-measure collection plans when handling is kept consistent.
Common research applications
- Quantify enzyfluo ampk phosphorylation in cells by Cell-Based ELISA (FL530/585 nm, 360/450 nm) readout.
- Compare treatment or phenotype groups using matched cells handling.
- Monitor time-course or pre/post changes in cells across study conditions.
Interpretation is usually strongest when signal changes are assessed alongside matrix-matched controls, replicate agreement, and the assay's stated analytical window.
Notes for experimental interpretation
- Matrix composition, background signal, and sample handling can influence apparent response; compare like-with-like whenever possible.
- Use appropriate blanks, controls, and replicate wells to distinguish biological differences from plate, reagent, or handling variability.
For laboratories requiring additional technical capacity, we provide scientific support services including assay execution, method guidance, product sourcing, and customization to align the assay with specific experimental objectives. If you need assistance selecting the appropriate kit configuration, adapting the workflow to your application, or identifying related research services, please click Talk to a Scientist, email support@biohippo.com, or review our Research Services; a member of our scientific team will follow up with recommendations tailored to your study.
6PGD Upregulation is Associated with Chemo-and Immuno-Resistance of Renal Cell Carcinoma via AMPK Signaling-Dependent NADPH-Mediated Metabolic Reprograming
Cao, J. et al (2020). 6PGD Upregulation is Associated with Chemo-and Immuno-Resistance of Renal Cell Carcinoma via AMPK Signaling-Dependent NADPH-Mediated Metabolic Reprograming. The American journal of the medical sciences, 360(3), 279-286. Assay: Phosphorylated AMPK in human renal carcinoma cells.
6PGD inhibition sensitizes hepatocellular carcinoma to chemotherapy via AMPK activation and metabolic reprogramming
Chen, Hu, et al (2019). 6PGD inhibition sensitizes hepatocellular carcinoma to chemotherapy via AMPK activation and metabolic reprogramming. Biomedicine & Pharmacotherapy 111: 1353-1358. Assay: AMPK phosphorylation in human hepatic cell.
“Trypsin-Treated beta-Lactoglobulin Improves Glucose Tolerance in C57BL/6 Mice by Enhancing AMPK Activation and Glucose Uptake in Hepatocytes
Tsuda, Yuichi, et al (2017). “Trypsin-Treated beta-Lactoglobulin Improves Glucose Tolerance in C57BL/6 Mice by Enhancing AMPK Activation and Glucose Uptake in Hepatocytes.” Biological and Pharmaceutical Bulletin 40.11: 1917-1922. Assay: AMPK phosphorylation in human hepatocytes.
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