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)
72-57-1 (tetrasodium salt) YesY
3D model (Jmol) Interactive image
ChemSpider 10482308 YesY
ECHA InfoCard 100.000.715
KEGG C19307 N
PubChem 5904246
Molar mass 872.88
Appearance deep blue in aqueous solution[1]
Melting point > 300 °C (572 °F; 573 K)
<0.1 mg/mL in water [2]
Solubility 20 mg/mL in methyl Cellosolve, and 0.6 mg/mL in ethanol
Lethal dose or concentration (LD, LC):
6200 mg/kg (oral, rat)
Except where otherwise noted, data are given for materials in their standard state (at 25 °C [77 °F], 100 kPa).
N verify (what is YesYN ?)
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. This dye may be a cause of certain birth defects such as encephalocele.

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.

The extinction coefficient for trypan blue is 6 x 104 M−1 cm−1 at 607nm in methanol.[3]

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.[4] The method cannot distinguish between necrotic and apoptotic cells.

It may be used to observe fungal hyphae[5] and stramenopiles.

Trypan blue is also used in ophthalmic cataract surgery to stain the anterior capsule in the presence of a mature cataract, to aid in visualization, before creating the continuous curvilinear capsulorhexis.

In early 20th century, the existence of a barrier protective toward the brain (blood brain barrier was inferred upon based on the observation that injection of trypan blue to animals led to whole‐body staining except for the brain and spinal cord.


Further reading


Wikimedia Commons has media related to Trypan blue staining.
  3. "Sigma-Aldrich, 60% Trypan Blue, Product page". Retrieved 2015-07-15.
  4. Strober, W (May 2001). "Trypan blue exclusion test of cell viability.". Current protocols in immunology. Appendix 3: Appendix 3B. doi:10.1002/0471142735.ima03bs21. PMID 18432654.
  5. Nowicki, Marcin; et al. (15 May 2013), A simple dual stain for detailed investigations of plant-fungal pathogen interactions, Vegetable Crops Research bulleting, InHort & Versita, doi:10.2478/v10032-012-0016-z, retrieved 2013-05-24
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