Measuring Cellular Metabolism Dynamics

Cells were seeded on a 96‐well plate with a density of 5 × 10⁴ cells/well and treated as indicated. Cells were then treated with ECAR reagents according to the manufacturer’s recommendations (Abcam). The ECAR measurements were taken at 5‐minute intervals for a total assay time of 120 minutes by a microplate reader system (PerkinElmer) using excitation and emission wavelengths of 380 and 615 nm, respectively. The OCR was measured in real time using the XF24 extracellular flux analyzer instrument (Seahorse Bioscience) as described previously.³⁵ (link), ³⁶ (link) Protein concentrations were used to normalize the results.

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Jiang D., Mo Q., Sun X., Wang X., Dong M., Zhang G., Chen F, & Zhao Q. (2021). Pyruvate dehydrogenase kinase 4‐mediated metabolic reprogramming is involved in rituximab resistance in diffuse large B‐cell lymphoma by affecting the expression of MS4A1/CD20. Cancer Science, 112(9), 3585-3597.

Publication 2021

XF24 extracellular flux analyzer instrument

Assay Cells Link protein Seahorse

Corresponding Organization : Qinghai Provincial Peoples Hospital

Other organizations : Guilin Medical University, Hainan Medical University

Top 5 similar protocols

Variable analysis

independent variables

Treatment

dependent variables

Extracellular Acidification Rate (ECAR)
Oxygen Consumption Rate (OCR)

control variables

Cell density (5 × 10^4 cells/well)
Assay time (120 minutes)
Measurement intervals (5 minutes)
Excitation and emission wavelengths (380 nm and 615 nm)
Protein concentrations (used to normalize results)

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