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Trypan Blue Stain (0.4%) - FAQs

Trypan Blue Stain (0.4%) - FAQs

View additional product information for Colorimetric Cell Viability Stains - FAQs (A40008024, T10282)

11 product FAQs found

Why am I getting a much higher bead count on the Countess 3 Standard Slide (Cat. No. A51876)?

When counting cells that have been mixed with Trypan Blue, the Countess accounts for this 1:1 dilution and multiplies the count by 2.
When using the Countess 3 Standard Slide, the Trypan Blue correction box needs to be unchecked, since a 1:1 dilution was not made with Trypan Blue.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Is there a limit to the number of times I can warm up Trypan Blue Stain (0.4%) for use with the Countess Automated Cell Counter (Cat. No. T10282)? What is the importance of warming up in aliquots of 1 mL?

The Trypan Blue Stain (0.4%) can be warmed up to 37 degrees C numerous times. The entire 1 mL volume of the stain should be warmed up to ensure that you are getting the entire amount at the proper concentration of 0.4%; if you took out an aliquot to warm up, and left behind some precipitate, the aliquot that is warmed up may not contain the complete amount of dye and the final concentration may be less than 0.4%.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

What is the shelf life of Trypan Blue Stain (0.4%) for use with the Countess Automated Cell Counter?

The shelf life of the product is 36 months from the date of production.

Find additional tips, troubleshooting help, and resources within our Cell Culture Support Center.

How long can cells be exposed to trypan blue before all cells are stained with the dye?

Unfortunately, there is no definitive answer to this question. The rate at which the cell-impermeant dye is absorbed depends on the cell type, their state of health, nourishment, engulfment activity, etc.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

What causes trypan blue to form a precipitate?

Exposure to light may degrade the dye and these contaminants may promote precipitation. Trypan blue can also form orange/red fibrous aggregates if exposed to refrigeration or freezing temperatures.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

How does trypan blue solution stain viable cells?

Trypan blue is a cell impermeant stain used to estimate the number of dead cells in a viable population. Its utility is based on the fact that it is a charged dye and does not enter cells unless the membrane is compromised. Live (viable) cells exclude the dye but dead (non-viable) cells or cells with a compromised membrane are stained an intense blue.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Prior to staining with trypan blue solution, do the cells need to be washed to remove the culture medium?

Trypan blue will bind to serum proteins as well as cellular proteins, which may result a high level of background staining. If the background is too dark, cells should be pelleted and resuspended in protein-free medium, buffer, or normal saline prior to counting.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Can I combine trypan blue staining with fluorescence staining?

It depends on the type of fluorescent stain used on the cells. Trypan blue is a cell-impermeant chromophore that can quench fluorescence. It may quench fluorescent staining on the surface of live cells or internal fluorescent staining in dead cells.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Can trypan blue solution be used to differentiate between apoptotic and necrotic cells?

Trypan blue will stain cells that have a compromised membrane. It cannot differentiate between compromised membranes caused by apoptosis or necrosis.

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

Do you have a protocol for cell counting via trypan blue exclusion?

Yes, you can find the protocol at the following link (https://www.thermofisher.com/us/en/home/references/gibco-cell-culture-basics/cell-culture-protocols/trypan-blue-exclusion.html)

Find additional tips, troubleshooting help, and resources within our Cell Analysis Support Center.

How much Trypan Blue solution should I add to my cell suspension before counting?

We recommend adding 0.1 mL of 0.4% Trypan Blue stock solution to 0.1 mL of cells. However, depending upon the sample and instrumentation, the ratio of 0.4% Trypan Blue solution to the volume of cells can be varied from 1: 10 to 1:1 (v/v) so that the final concentration of Trypan Blue is 0.04% - 0.2%.

Find additional tips, troubleshooting help, and resources within our Cell Counting Support Center.