basic fuchsin-methylene blue stain

Methylene blue

Methylene blue is a heterocyclic aromatic chemical compound with molecular formula: C16H18ClN3S. It has many uses in a range of different fields, such as biology or chemistry. At room temperature it appears as a solid, odorless, dark green powder, that yields a blue solution when dissolved in water. The hydrated form has 3 molecules of water per molecule of MB. Methylene blue should not be confused with methyl blue, another histology stain, new methylene blue, nor with the methyl violets often used as pH indicators.



Methylene blue is widely used as a redox indicator in analytical chemistry. Solutions of this substance are blue when in an oxidizing environment, but will turn colorless if exposed to a reducing agent. The redox properties can be seen in a classical demonstration of chemical kinetics in general chemistry, the "blue bottle" experiment. Typically, a solution is made of dextrose, methylene blue, and sodium hydroxide. Upon shaking the bottle, oxygen oxidizes methylene blue, and the solution turns blue. The dextrose will gradually reduce the methylene blue to its colorless, reduced form. Hence, when the dissolved oxygen is entirely consumed, the solution will turn colorless.

Methylene blue is also a photosensitizer used to create singlet oxygen when exposed to both oxygen and light. It is used in this regard to make organic peroxides by a Diels-Alder reaction which is spin forbidden with normal atmospheric triplet oxygen.


In biology methylene blue is used as a dye for a number of different staining procedures, such as Wright's stain and Jenner's stain. Since it is a temporary staining technique, methylene blue can also be used to examine RNA or DNA under the microscope or in a gel: as an example, a solution of methylene blue can be used to stain RNA on hybridization membranes in northern blotting to verify the amount of nucleic acid present. While methylene blue is not as sensitive as ethidium bromide, it is less toxic and it does not intercalate in nucleic acid chains, thus avoiding interference with nucleic acid retention on hybridization membranes or with the hybridization process itself.

It can also be used as an indicator to determine if a cell such as yeast is alive or not. The blue indicator turns colorless in the presence of active enzymes, thus indicating living cells. However if it stays blue it doesn't mean that the cell is dead - the enzymes could be inactive/denatured. It must be noted that methylene blue can inhibit the respiration of the yeast as it picks up hydrogen ions made during the process. The yeast cell cannot then use those ions to release energy.

In neuroscience, methylene blue can also serve as a non-selective inhibitor of NO synthase.



Methylene blue was identified by Paul Ehrlich about 1891 as a successful treatment for malaria. It disappeared as an anti-malarial during the Pacific War in the tropics, since American and Allied soldiers disliked its two prominent, but reversible side effects: tuning the urine green, and the sclera (the whites of the eyes) blue. Interest in its use a an anti-malarial has recently been revived, especially because it is very cheap. Several clinical trials are in progress, trying to find a suitable drug combination. Initial attempts to combine methylene blue with chloroquine were disappointing; however, more recent attempts have appeared more promising.

Owing to its reducing agent properties, methylene blue is employed as a medication for the treatment of methemoglobinemia, which can arise from ingestion of certain pharmaceuticals or broad beans. Basically, methylene blue acts to reduce the heme group from methemoglobin to hemoglobin. Methylene blue also blocks accumulation of cyclic guanosine monophosphate (cGMP) by inhibiting the enzyme guanylate cyclase: this action results in reduced responsiveness of vessels to cGMP-dependent vasodilators like nitric oxide and carbon monoxide.

Methylene blue is used in endoscopic polypectomy as an adjunct to saline or epinephrine, and is used for injection into the submucosa around the polyp to be removed. This allows the submucosal tissue plane to be identified after the polyp is removed, which is useful in determining if more tissue needs to be removed, or if there has been a high risk for perforation. Methylene blue is also used as a dye in chromoendoscopy, and is sprayed onto the mucosa of the gastrointestinal tract in order to identify dysplasia, or pre-cancerous lesions. Intravenously injected methylene blue is readly released into the urine and thus can be used to test the urinary tract for leaks or fistulas.

In surgeries such as sentinel lymph node dissections, methylene blue can be used to visually trace the lymphatic drainage of pertinent tissues. Similarly, methylene blue is added to bone cement in orthopedic operations to provide easy discrimination between native bone and cement. Additionally, methylene blue accelerates the hardening of bone cement, increasing the speed at which bone cement can be effectively applied.

Poisoning : Since its reduction potential is similar to that of oxygen and can be reduced by components of the electron transport chain, large doses of methylene blue are sometimes used as an antidote to potassium cyanide poisoning, a method first successfully tested in 1933 by Dr. Matilda Moldenhauer Brooks in San Francisco. Methylene blue was also used at mid-century in the treatment of carbon monoxide poisoning.

Methylene blue is a monoamine oxidase inhibitor, and if infused intravenously at doses exceeding 5 mg/kg, may precipitate serious serotonin toxicity, serotonin syndrome, if combined with any selective serotonin reuptake inhibitors (SSRIs) or other serotonin reuptake inhibitor (e.g., duloxetine, sibutramine, venlafaxine, clomipramine, imipramine).

Methylene blue has been used as a placebo .

Ifosfamide neurotoxicity

Another, less well-known use of methylene blue is its utility for treating ifosfamide neurotoxicity. Methylene blue was first reported for treatment and prophylaxis of ifosfamide neuropsychiatric toxicity in 1994. A toxic metabolite of ifosfamide, chloracetaldehyde (CAA), disrupts the mitochondrial respiratory chain, leading to an accumulation of nicotinamide adenine dinucleotide hydrogen (NADH). Methylene blue acts as an alternative electron acceptor, and reverses the NADH inhibition of hepatic gluconeogenesis while also inhibiting the transformation of chloroethylamine into chloroacetaldehyde, and inhibits multiple amine oxidase activities, preventing the formation of CAA The dosing of methylene blue for treatment of ifosfamide neurotoxicity varies, depending upon its use simultaneously as an adjuvant in ifosfamide infusion, versus its use to reverse psychiatric symptoms that manifest after completion of an ifosfamide infusion. Reports suggest that methylene blue at 50-60mg up to six doses a day have resulted in improvement of symptoms within 10 minutes to several days.. Alternatively, it has been suggested that intravenous methylene blue 50mg every six hours for prophylaxis during ifosfamide treatment in patients with history of ifosfamide neuropsychiatric toxicity. Prophylactic administration of 50mg of methylene blue the day before initiation of ifosfamide, and 50mg three times daily during ifosfamide chemotherapy has been recommended to lower the occurrence of ifosfamide neurotoxicity.

Clinical trials

TauRx Therapeutics has reported that methylene blue (methylthioninium chloride), under the tradename rember, may provide a way of halting or slowing the progression of Alzheimer's dementia. However, the formulation used was different from that commonly available as a medicine and caution has been expressed about use of methylene blue as a treatment for Alzheimer's. TauRx Therapeutics has suggested that the mechanism by which methylene blue might delay or reverse neurodegeneration in Alzheimer's disease is as an inhibitor of Tau protein aggregation. While methylene blue arguably has an effect on Tau aggregation, it also has an effect on mitochondrial function which is likely to play an important role. In vitro studies suggest that methylene blue might be an effective remedy for both Alzheimer's and Parkinson's disease by enhancing key mitochondrial biochemical pathways. It can disinhibit and increase complex IV, whose inhibition correlates with Alzheimer's disease.

Methylene blue might also delay senescence as one study has shown that it extended the lifespan of IMR90 fibroblasts by more than 20 population doublings.


Methylene blue is used in aquaculture and by tropical fish hobbyists as a treatment for fungal infections. It can also be effective in treating fish infected with ich, the parasitic protozoa Ichthyophthirius multifiliis. It is usually used to protect newly laid fish eggs from being infected by fungus or bacteria. This is useful when the hobbyist wants to artificially hatch the fish eggs.

As a Prank on TV

An episode of M*A*S*H, "Sons and Bowlers", showed Major Winchester using a dose of methylene blue to take down a rival camp's bowling champion during a contest. The champ panics when his urine turns blue, and listens to Winchester's advice to refrain from all exercise – including bowling, which allows the 4077th to win. In an episode of "ER" from the 5th season they use it to play an April Fools' Day prank on the desk clerk Jerry. Small methylene blue amounts cause faint green or blue colors; larger amounts cause a deeper blue color. It is not known if this prank is realistic because of the bitterness and color that will be added to the drink.

Adverse Reactions

Cardiovascular Central Nervous System Dematologic Gastrointestinal Genitourinary Hematologic
•Precordial pain
•Mental confusion
•Staining of skin
•Injection site necrosis (SC)
•Fecal discoloration
•Abdominal pain
•Discoloration of urine
•Bladder irritation


See also

External links

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