Trypan blue is an azo dye. It is a direct dye for cotton textiles. In biosciences, it is used as a

vital stain A vital stain in a casual usage may mean a stain that can be applied on living cells without killing them. Vital stains have been useful for diagnostic and surgical techniques in a variety of medical specialties. In supravital staining, living cells ...

to selectively colour dead tissues or cells blue. Live cells or tissues with intact

cell membrane The cell membrane (also known as the plasma membrane (PM) or cytoplasmic membrane, and historically referred to as the plasmalemma) is a biological membrane that separates and protects the interior of all cells from the outside environment ( ...

s 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 appear 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 and chemistry

Trypan blue is derived from

toluidine There are three isomers of toluidine, which are organic compounds. These isomers are ''o''-toluidine, ''m''-toluidine, and ''p''-toluidine, with the prefixed letter abbreviating, respectively, ''ortho''; ''meta''; and ''para''. All three are ary ...

, that is, any of several isomeric bases, C₁₄H₁₆N₂, derived from

toluene Toluene (), also known as toluol (), is a substituted aromatic hydrocarbon. It is a colorless, water-insoluble liquid with the smell associated with paint thinners. It is a mono-substituted benzene derivative, consisting of a methyl group (CH3) a ...

. Trypan blue is so-called because it can kill

trypanosomes Trypanosomatida is a group of kinetoplastid excavates distinguished by having only a single flagellum. The name is derived from the Greek ''trypano'' (borer) and ''soma'' (body) because of the corkscrew-like motion of some trypanosomatid species. ...

, the parasites that cause

sleeping sickness African trypanosomiasis, also known as African sleeping sickness or simply sleeping sickness, is an insect-borne parasitic infection of humans and other animals. It is caused by the species ''Trypanosoma brucei''. Humans are infected by two typ ...

. An analog of trypan blue,

suramin Suramin is a medication used to treat African sleeping sickness and river blindness. It is the treatment of choice for sleeping sickness without central nervous system involvement. It is given by injection into a vein. Suramin causes a fair ...

, is used pharmacologically against

trypanosomiasis Trypanosomiasis or trypanosomosis is the name of several diseases in vertebrates caused by parasitic protozoan trypanosomes of the genus ''Trypanosoma''. In humans this includes African trypanosomiasis and Chagas disease. A number of other diseas ...

. Trypan blue is also known as diamine blue and Niagara blue. The extinction coefficient for trypan blue is 6⋅10⁴ M⁻¹ cm⁻¹ at 607 nm in methanol. Trypan red and trypan blue were first synthesized by the German scientist

Paul Ehrlich Paul Ehrlich (; 14 March 1854 – 20 August 1915) was a Nobel Prize-winning German physician and scientist who worked in the fields of hematology, immunology, and antimicrobial chemotherapy. Among his foremost achievements were finding a cure ...

in 1904.

Uses of trypan blue

Trypan blue is commonly used in microscopy (for cell counting) and in

laboratory mice The laboratory mouse or lab mouse is a small mammal of the order Rodentia which is bred and used for scientific research or feeders for certain pets. Laboratory mice are usually of the species ''Mus musculus''. They are the most commonly used ...

for assessment of tissue viability. The method cannot distinguish between

necrotic Necrosis () is a form of cell injury which results in the premature death of cells in living tissue by autolysis. Necrosis is caused by factors external to the cell or tissue, such as infection, or trauma which result in the unregulated dige ...

and

apoptotic Apoptosis (from grc, ἀπόπτωσις, apóptōsis, 'falling off') is a form of programmed cell death that occurs in multicellular organisms. Biochemical events lead to characteristic cell changes ( morphology) and death. These changes incl ...

cells. It may be used to observe fungal hyphae and

stramenopiles Stramenopile is a clade of organisms distinguished by the presence of stiff tripartite external hairs. In most species, the hairs are attached to flagella, in some they are attached to other areas of the cellular surface, and in some they have be ...

Vesicular Arbuscular Mycorrhizae 40X0031 03

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 keratoplasty, trypan blue can be used to stain the posterior stromal fibers during deep lamellar endothelial keratoplasty (DLEK) and to stain the endothelium in Descemet's stripping endothelial keratoplasty (DSEK). Trypan blue is used in vitreoretinal surgeries also. In early 20th century, the existence of a barrier protective toward the brain (

blood brain barrier Blood is a body fluid in the circulatory system of humans and other vertebrates that delivers necessary substances such as nutrients and oxygen to the cells, and transports metabolic waste products away from those same cells. Blood in the c ...

) was inferred, based on the observation that injection of trypan blue in animals led to whole‐body staining except for the brain and spinal cord.

Synonyms

*Azidine blue 3B *Benzamine blue 3B *Benzo Blue bB *Chlorazol blue 3B *Diamine blue 3B *Dianil blue H3G *Direct blue 14 *Niagara blue 3B

References

{{Glutamate metabolism and transport modulators Azo dyes Cell culture reagents Vital stains IARC Group 2B carcinogens Naphthalenesulfonic acids 1-Naphthols Naphthylamines fr:Bleu de trypan#Coloration au bleu de trypan

Background and chemistry

Uses of trypan blue

Synonyms

Further reading

References