Babco, Signet, Sternberger Monoclonals, Senetek, Covance

other

Zombie R718™ Fixable Viability Kit

423115

100 tests

89 USD

423115

Zombie R718™ Fixable Viability Kit

100 tests

89 USD

NONAB_CHMPRB

Zombie R718™ dye is excited by the red laser (633 nm) and has fluorescence emission maximum at 718 nm. If using in a multi-color panel design, filter optimization may be required depending on other fluorophores used. Zombie R718™ dye has similar spectra to Spark Red™ 718.Standard Cell Staining Protocol:;Prior to reconstitution, spin down the vial of lyophilized reagent in a microcentrofuge to ensure the reagent is at the bottom of the vial.;For reconstitution, pre-warm the kit to room temperature; add 100 µL of DMSO to one vial of Zombie R718™ dye and mix until fully dissolved.;Wash cells with PBS buffer (no Tris buffer and protein free).;Dilute Zombie R718™ dye at 1:100-1000 in PBS. Resuspend 1-10 x 10^6 cells in diluted 100 µL Zombie R718™ solution. To minimize background staining of live cells, titrate the amount of dye and/or number of cells per 100 µL for optimal performance. Different cell types can have a wide degree of variability in staining based on cell size and degree of cell death.Note: Don't use Tris buffer as a diluent and be sure that the PBS does not contain any other protein like BSA or FBS.;Note: The amount of dye used can also influence the ability to detect apoptotic as well as live and dead cells.;Incubate the cells at room temperature (or 4°C), in the dark, for 15-30 minutes.;Wash one time with 2 mL BioLegend's Cell Staining Buffer (Cat. No. 420201) or equivalent buffer containing serum or BSA.;Continue performing antibody staining procedure as desired.;Cells can be fixed with paraformaldehyde or methanol prior to permeabilization or can be analyzed without fixation.;No-wash Sequential Staining Protocol:;Wash cells with PBS buffer (no Tris buffer and protein free).;For reconstitution, pre-warm the kit to room temperature; add 100 µL of DMSO to one vial of Zombie R718™ dye and mix until fully dissolved.;Determine the total µL volume of antibody cocktail previously titrated and optimized for the assay that will be added to each vial/well of cells based on a final volume of 100 µL. Subtract that antibody volume from the 100 µL total staining volume intended for the assay. In the remaining volume, dilute Zombie R718™ dye at 1:100-1000 in PBS as determined by prior optimization at that volume. For example, if you are adding 20 µL of antibody cocktail for a 100 µL total staining volume, use 80 µL of Zombie R718™ solution. Resuspend 1-10 x 10^6 cells in the appropriate volume of Zombie R718™ solution. Different cell types can have a wide degree of variability in staining based on cell size and degree of cell death.Note: Don't use Tris buffer as a diluent and be sure that the PBS does not contain any other protein like BSA or FBS.;Note: The amount of dye used can also influence the ability to detect apoptotic as well as live and dead cells.;Incubate for 10-15 minutes at RT (or 4°C), protected from light. Without washing the cells, add the cell surface antibody cocktail and incubate for another 15-20 minutes.;Add 1-2 mL Cell Staining Buffer (Cat. No. 420201) or equivalent buffer containing BSA or serum. Centrifuge to pellet.;Continue with normal fixation and permeabilization procedure. If planning to skip fixation and analyze cells live, complete an additional wash step to minimize any unnecessary background of the live cells.Notes: If the cell type in use cannot tolerate a protein-free environment, then titrate the Zombie R718™ dye in the presence of the same amount of BSA/serum as will be present in the antibody staining procedure. A higher amount of Zombie R718™ may be required since the BSA/serum will react with and bind up some proportion of the Zombie R718™.;