Improve Research Reproducibility A Bio-protocol resource
Concise Method
tooltip

Measurement of glycolysis as extracellular acidification rate

YJ Yan Jouroukhin
YK Yusuke Kageyama
VM Varvara Misheneva
AS Alexey Shevelkin
SA Shaida Andrabi
EP Emese Prandovszky
RY Robert H. Yolken
VD Valina L. Dawson
TD Ted M. Dawson
SA Susan Aja
HS Hiromi Sesaki
MP Mikhail V. Pletnikov
ask Ask a question
Favorite

Primary astrocytes were plated at a density of 50 × 103 per well in XF24 cell-culture microplates for 48 h. On the day of assay, the medium was replaced with glucose-free XF24 Seahorse medium. Glycolytic flux (basal glycolysis, glycolytic capacity, and glycolytic reserve) was assessed as the extracellular acidification rate (ECAR) by sequentially adding glucose, oligomycin, and 2-deoxyglucose to the XF24 flux analyzer23. ECAR was measured at 37 °C with a 1-min mix, 1-min wait, and 2-min measurement protocol. Seahorse analysis was started after 45 min of incubation in a CO2-free incubator. The first ECAR measurement was recorded after 11-min equilibration, a 1-min mix period, and a 1-min wait period.

Copyright and License information: The Author(s) ©2018
Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

Do you have any questions about this protocol?

Post your question to gather feedback from the community. We will also invite the authors of this article to respond.

post Post a Question
0 Q&A