Fixable Viability Dyes

Retain fluorescence performance to discriminate live and dead cells without any loss of staining intensity post fixation or permeabilization. 

Figure 1. Comparison of fluorescent staining with iQue^® Fixable Viability Dyes in fixed/permeabilized cells and non-fixed cells. 
A mixture of viable and heat-treated (65°C, 10 minutes) non-viable Jurkat cells were stained with iQue^® Fixable V/Blue (A), B/Green (B), or R/Red (C) dye. The stained cells were either unfixed or fixed and permeabilized with 3.7% formaldehyde, 0.1% saponin in PBS. All the samples were acquired with an iQue^® HTS instrument and histograms were derived from gated singlets using the  pre-set templates included in the kit.

Analyze surface and intracellular marker expression simultaneously while minimizing the potential artifacts from dead cells.

Figure 2. Example of Detection and Analysis of intracellular IFN-γ expression in iQue^® Fixable Viability Dye-stained PBMCs.
PBMCs were cultured in CD3 antibody-coated plate for two days in the presence of 10 ng/mL human recombinant IL-2. Protein transport inhibitor monensin (2 µM) was added to the cell culture 4 hours before the assay. Activated PBMCs (20 µL) were  added to each well of a 96-well assay plate, stained with iQue^® Fixable Viability (V/Blue) Dye, followed by staining with T cell surface marker CD3. The cells were then fixed and permeabilized with Cytofix/Cytoperm solution (BD Biosciences), stained with PE-conjugated mouse anti-human IFN-γ antibody. The samples were acquired with an iQue^® HTS instrument and data were analyzed with iQue Forecyt^® Software. A, Gate of all cells. B, Gate of singlets. C, Exclusion of dead cells with iQue^® Fixable Viability (V/Blue) Dye staining. D, Plot of IFN-γ expression in CD3+ live cells.

Conserve samples to accommodate multiple down stream assays with only 20 µL of cells required.

Figure 3. Easy to follow, low volume (20 µL/well) protocol save cells and setup time