NIR-Red Dead Cell-1 Dye

NIR-Red Dead Cell-1 Dye (100 nM; 15 min) can accurately distinguish necrotic cells (binding to both) from live cells (showing only red autofluorescence) in flow cytometry experiments of Jurkat and HT-29 cells when co-stained with Annexin V-Cy5.5^[1].
NIR-Red Dead Cell-1 Dye (64.5 μg/mL; 15 min) colocalizes with DAPI in confocal microscopy experiments and binds to nuclear DNA specifically labeled with green fluorescence in Jurkat cells treated with Camptothecin (HY-16560)^[1].
NIR-Red Dead Cell-1 Dye (100 nM; 15 min) reflects cell death more accurately than 5-CFDA-AM (HY-131131) in viability assays of algal cells such as Brachiomonas submarina and Tetraselmis suecica^[2].
NIR-Red Dead Cell-1 Dye can be used to directly measure live and dead cells adhering to diatoms^[3]. TO-PRO-1 can only stain dead cells. When NIR-Red Dead Cell-1 Dye was used to stain the marine diatom Nitzchia closterium, live and dead cells were identified as red and yellow, respectively, under a blue-excited epifluorescence microscope. Live cells appear red due to the autofluorescence of intracellular chlorophyll, while dead cells appear yellow due to the fluorescence of TO-PRO-1^[3].

Guide (The following is our recommended solution. This solution is merely a guideline and should be modified according to your specific needs.)
1. Cell Preparation
The cells (such as Jurkat, HT-29 or algal cells) are suspended in phosphate-buffered saline (PBS) at a density of 1×10⁵-1×10⁶ cells/mL.
Induce cell necrosis by treating with Camptothecin (HY-16560) (5 μM, 10 h), 5-FU (HY-90006)/Oxaliplatin (HY-17371), or γ-irradiation (lethal dose: 2400-4800 Gy).
2. Staining Protocol
Dye Preparation: Prepare the 1 mM DMSO stock solution of NIR-Red Dead Cell-1 Dye and dilute it to 100 nM (for mammalian cells) or 64.5 μg/mL (for algal cells) using PBS.
Sample Incubation: Add 5 μL of the dye to 30 μL of the cell suspension, mix gently, and incubate at room temperature in the dark for 15 minutes.
Cleaning (for algal cells): Centrifuge at 2,500 g for 2 minutes, discard the supernatant, and resuspend in fresh PBS to reduce background fluorescence.
3. Detection Method
① Fluorescence Microscopy Method: Use a confocal fluorescence microscope with a blue filter (excitation: 400-550 nm, emission: 531 nm) to detect the green fluorescence of NIR-Red Dead Cell-1 Dye.
Count at least 300 cells, and distinguish between dead cells (bright green cell nucleus) and live cells (red chlorophyll autofluorescence in algae or no green fluorescence in mammalian cells).
② Flow Cytometry Method: Use 488 nm excitation light to detect the emission at 530-550 nm.
Set gates based on forward/side scatter and fluorescence intensity to quantify the necrotic cell population.
4. Control Settings
Positive Control: 100% live cells (untreated, expected low binding rate of NIR-Red Dead Cell-1 Dye).
Negative Control: 100% non-living cells (radiation killed, expected high binding rate of NIR-Red Dead Cell-1 Dye).

MedChemExpress (MCE) has not independently confirmed the accuracy of these methods. They are for reference only.