Protocol Type: in vitro assay
Protocol Name: NCL Method ITA-7: Detection of Nitric Oxide Production by the Macrophage Cell Line RAW264.7
Protocol Abbreviation: ITA-7
Protocol Version: 2
DOI: http://dx.doi.org/10.17917/T6JC-ZG30
Protocol File: 20240411_12-02-35-338_NCL_Method_ITA-7.pdf
File Title: ITA-7
Description: This document describes a protocol for quantitative determination of nitrite (NO2-), a stable oxidative end-product of the antimicrobial effector molecule nitric oxide in cell culture medium. The protocol is used to evaluate the capability of nanomaterials to induce nitric oxide production by macrophages. Nitric oxide secreted by macrophages has a half-life of seconds and interacts with a number of different molecular targets, resulting in cytotoxicity. In the presence of oxygen and water, nitric oxide interacts with itself to generate other reactive nitrogen oxide intermediates and ultimately decomposes to form nitrite (NO2‚àí) and nitrate (NO3‚àí). Interestingly, despite expression of inducible nitric oxide synthase (iNOS), an enzyme responsible for the generation of nitric oxide in both human and mouse immune cells, the induction of nitric oxide by immunologically active agonists (e.g., bacterial lipopolysaccharide) and the levels of produced nitric oxide are different between human and mouse immune cells. The response in human immune cells, especially under in vitro conditions is substantially lower than in murine cells. For this reason, a murine macrophage cell line is a better model for in vitro analysis of nitric oxide production than human monocytes and macrophages.
Resource Type: Protocol
Data Access: https://general.datacommons.cancer.gov/#/data