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Application note: Hypoxia measurements in live and fixed cells

Hypoxia measurements in live and fixed cells using fluorescence microscopy and high-content imaging

Hypoxia is an important phenomenon in many physiological processes and involved in many human diseases including cancers, cardiovascular diseases, and neurodegenerative diseases.

The study of hypoxia has been complicated by the lack of sensitive dyes for measuring hypoxia at greater than 1% O2. Here we describe a live cell–based method to conveniently measure hypoxia using Invitrogen™ ImageiT ™ Green Hypoxia Reagent. The probe is sensitive to varying concentrations of oxygen and can detect O2 concentrations as low as 5% in cells. Using this probe, we measured hypoxia in several cell lines, including A549, HeLa, and U2OS, using fluorescence microscopy, high-content imaging, and a fluorescence plate reader. Image-iT Green Hypoxia Reagent can be multiplexed with indicators for other important physiological parameters like mitochondrial membrane potential, apoptosis, and oxidative stress.

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