Quantitative Nitric Oxide Measurement

Nitric oxide (NO) release into the cell culture media was measured with a Nitric Oxide Analyzer (Sievers 270B) according to lab standards^{94 (link)}. The system measures nitrite, the major product of NO, which is generated when NO is exposed to superoxide anion. Therefore, NO that is produced in the media is able to be measured as nitrite after exposure to iodide and acetic acid at room temperature. Ionomycin from streptomyces conglobatus (Sigma) was added 1 h before collection to serve as a positive control. The media was collected for NO measures, and the cells were collected in protein lysis buffer to measure the protein level. All NO measurements were corrected for total protein (mg) based on whole cell lysates.

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Zhou W., Li W., Wang S., Salovska B., Hu Z., Tao B., Di Y., Punyamurtula U., Turk B.E., Sessa W.C, & Liu Y. (2023). An optogenetic-phosphoproteomic study reveals dynamic Akt1 signaling profiles in endothelial cells. Nature Communications, 14, 3803.

Publication 2023

Ionomycin from streptomyces conglobatus

Acetic acid Buffer Cell Cell culture Culture media Iodide Ionomycin Nitric oxide Nitrite Protein Streptomyces conglobatus Superoxide anion

Corresponding Organization :

Other organizations : Yale University, Sichuan University, Brown University, Providence College, Yale Cancer Center

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Variable analysis

independent variables

Ionomycin from streptomyces conglobatus (Sigma) was added 1 h before collection

dependent variables

Nitric oxide (NO) release into the cell culture media
Nitrite, the major product of NO
Protein level in the cells

control variables

Nitrite was generated when NO was exposed to superoxide anion
Nitrite was measured after exposure to iodide and acetic acid at room temperature
All NO measurements were corrected for total protein (mg) based on whole cell lysates

positive control

Ionomycin from streptomyces conglobatus (Sigma) was added 1 h before collection to serve as a positive control

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