1.Zhang, S., Hosaka, M., Yoshihara, T., Negishi, K., Iida, Y., Tobita, S., & Takeuchi, T. (2010) 磷光–发射铱复合物作为一个缺氧感应探头,用于活体动物中的肿瘤成像。Cancer Research 70(11), 4490-4498.
Can Image-iT Red Hypoxia Reagent be used in cells expressing fluorescent proteins?
Yes, as long as the fluorescent protein does not emit in the red range (610 nm). For cells that do not express a fluorescent protein, we recommend viewing unstained cells under the TRITC and Texas Red channels to examine autofluorescence. For cells expressing a fluorescent protein, we recommend analyzing unstained cell samples to determine the extent the fluorescent protein emission may overlap in these channels.
May I include a total cell stain (to stain all cells) along with Image-iT Hypoxia Reagent?
You may use cell-permeable nuclear counterstains or surface labels, but avoid any general cytoplasmic stains that may interfere with the emission of the hypoxia reagents.
May I include a dead cell stain along with Image-iT Hypoxia Reagent?
Yes. We recommend using NucGreen Dead 488 ReadyProbes Reagent (Cat No. R37109) with Image-iT Red Hypoxia Reagent and NucRed Dead 647 ReadyProbes Reagent (Cat No. R37113) with Image-iT Green Hypoxia Reagent.
Which filter set can I use for optimal excitation and emission with Image-iT Red Hypoxia Reagent?
The Image-iT Red Hypoxia Reagent has an approximate Ex/Em maxima at 490/610 nm. You may use a 488 nm laser and a Texas Red emission filter. For lamp-based instruments, a YFP longpass filter set may be used (excitation range from 485 to 510 nm and emission from 550 nm up to 620 nm).