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Intracellular Nitric Oxide (NO) Assays
Altered NO production is implicated in various immunological, cardiovascular, neurodegenerative and inflammatory diseases. As a free radical, NO is rapidly oxidized and exists in relatively low concentration. It has been challenging to detect and understand the role of NO in biological systems. Cell Meter™ Fluorimetric Intracellular Nitric Oxide Assay Kits provide a robust tool to monitor intracellular NO level in live cells.
Fig. 1
Detection of exogenous nitric oxide (NO) in cells upon DEA NONOate treatment (NO donor) using Cell Meter™ Fluorimetric Intracellular Nitric Oxide Assay Kit (Cat#16350). CHO-K1 and HeLa cells at 50,000 cells/well/100 µL were seeded overnight in a 96-well black wall/clear bottom plate. Cells were incubated with Nitrixyte™ Orange working solution at 37 °C for 30 minutes. The cells were treated with or without 1mM DEA NONOate at 37 °C for 30 minutes. The fluorescence signal was monitored at Ex/Em = 540/590 nm (cut off = 570 nm) with bottom read mode.
Nitrixyte™ Orange and Nitrixyte™ Red are developed as excellent replacements for DAF-2 for the detection and imaging of free NO in cells. Compared to the widely used DAF-2 probes, Nitrixyte™ Orange and Nitrixyte™ Red have better photostability and enhanced cell permeability. Cell Meter™ Fluorimetric Intracellular Nitric Oxide Assay Kits (Cat# 16350 & 16351) use Nitrixyte™ Orange that reacts with NO to generate a bright orange fluorescent product. The NO-generated product of Nitrixyte™ Orange has spectral properties similar to those of Cy3® and TRITC. Nitrixyte™ Orange can be readily loaded into live cells, and its fluorescence signal can be conveniently monitored using the filter set of Cy3® or TRITC. Kit 16350 is optimized for fluorescence imaging and microplate reader applications. Kit 16351 is optimized for flow cytometry applications.
Fig. 2
Fluorescence images of endogenous nitric oxide (NO) measurement in RAW 264.7 macrophage cells using Cell Meter™ Fluorimetric Intracellular Nitric Oxide Assay Kit (Cat#16350). Raw 264.7 cells at 100,000 cells/well/100 µL were seeded overnight in a 96-well black wall/clear bottom plate. Cells were incubated with Nitrixyte™ Orange, and treated with (Right) or without (Left) 20 µg/mL of lipopolysaccharide (LPS) and 1 mM L-Arginine (L-Arg) at 37 °C for 16 hours.
Fig. 3
Detection of exogenous nitric oxide (NO) in Jurkat cells upon DEA NONOate treatment (NO donor) using Cell Meter™ Fluorimetric Intracellular Nitric Oxide Assay Kit (Cat#16351). Cells were incubated with Nitrixyte™ Orange at 37 °C for 30 minutes and washed with assay buffer twice. The cells were treated with (Red) or without (Blue) 1mM DEA NONOate at 37 °C for 30 minutes.
Cell Meter™ Fluorimetric Intracellular Nitric Oxide Assay Kit 16356 uses Nitrixyte™ Red that reacts with NO to generate a bright red fluorescent product. The NO-generated fluorescent product of Nitrixyte™ Red has spectral properties similar to those of Texas Red®. Nitrixyte™ Red can be readily loaded into live cells, and its fluorescence signal can be conveniently monitored using the filter set of Texas Red®. This kit is optimized for flow cytometry applications.
Document: 03.0064.141001r1
Last updated Tue Sep 09 2025
Intracellular Nitric Oxide (NO) Assays