Trypan blue

Trypan Blue
IUPAC name	(3Z,3'Z)-3,3'-[(3,3'-dimethylbiphenyl-4,4'-diyl)di(1Z)hydrazin-2-yl-1-ylidene]bis(5-amino-4-oxo-3,4-dihydronaphthalene-2,7-disulfonic acid)
Identifiers
CAS number	72-57-1 N,(tetrasodium salt)
PubChem	5904246
SMILES	CC1=C(C=CC(=C1)C2=CC(=C(C=C2)N/N=C/3\C(=CC4=CC(=CC (=C4C3=O)N)S(=O)(=O)O)S(=O)(=O)O)C)N/N=C/5\C(=CC6=CC (=CC(=C6C5=O)N)S(=O)(=O)O)S(=O)(=O)O
Properties
Molecular formula	C34H28N6O14S4
Molar mass	872.88 g mol−1
N(what is this?)  (verify) Except where noted otherwise, data are given for materials in their standard state (at 25 °C, 100 kPa)
Infobox references

Trypan blue is a vital stain used to selectively colour dead tissues or cells blue. It is a diazo dye.

Live cells or tissues with intact cell membranes are not coloured. Since cells are very selective in the compounds that pass through the membrane, in a viable cell Trypan blue is not absorbed; however, it traverses the membrane in a dead cell. Hence, dead cells are shown as a distinctive blue colour under a microscope. Since live cells are excluded from staining, this staining method is also described as a Dye Exclusion Method.

Background & chemistry

Trypan blue is derived from toluidine, that is, any of several isomeric bases, C₁₄H₁₆N₂, derived from toluene. Trypan blue is so-called because it can kill trypanosomes, the parasites that cause sleeping sickness. An analog of trypan blue, suramin is used pharmacologically against trypanosomiasis. Trypan blue is also known as diamine blue and Niagara blue.

Trypan red and Trypan blue were first synthesized by the German scientist Paul Ehrlich in 1904.

Uses of trypan blue

Observation with an optical microscope of Hyaloperonospora parasitica within a leaf of Arabidopsis thaliana by using the trypan blue staining.

Trypan blue is commonly used in microscopy (for cell counting) and in laboratory mice for assessment of tissue viability. The method cannot distinguish between necrotic and apoptotic cells.

It may also be used to observe fungal hyphae and Stramenopiles.

Protocol for vital staining

Place a suitable volume of a cell suspension (20-200 μL) in appropriate tube add an equal volume of 0.4% Trypan blue and gently mix, let stand for 5 minutes at room temperature. Place 10 μl of stained cells in a Hemocytometer and count the number of viable (unstained) and dead (stained) cells. Calculate the average number of unstained cells in each quadrant, and multiply by 2*10⁴ to find the number of viable cells/ml. The percentage of viable cells is the [number of viable cells] divided by the total number of [dead plus viable cells], multiplied by 100. % viable cells = 100 x [live cells]/[dead + live cells]

References

• Chapter "Detection of Caspase Activation Combined with Other Probes of Apoptosis", Eurekah Bioscience Collection, NCBI bookshelf
• Protocol for use of the dye from Northwestern University (PDF)

Synonyms

• Azidine Blue 3B
• Benzamine Blue 3B
• Benzo Blue 3B
• Chlorazol Blue 3B
• Diamine Blue 3B
• Dianil Blue H3G
• Direct Blue 14
• Niagara Blue 3B

External links

Wikimedia Commons has media related to: Trypan_blue_staining