Supra vital staining in early diagnosis of mucosal malignancies

From Otolaryngology Online


Supravital staining of suspicious mucosal lesions in oral cavity helps in the early diagnosis of malignant lesions. The dye used for this purpose is usually acidophilic in nature and is known to preferentially stain acidic cellular components. At this juncture it should be pointed out that DNA & RNA are acidic in nature.


Toludine blue is the dye which is preferred for the purpose of supravital staining. This dye is acidophilic in nature and preferentially stains acidic cellular components like DNA & RNA. Dysplastic mucosa / Malignant lesions of mucosa have large amounts of acidic DNA & RNA due to the active cellular proliferation which is occurring there. In this test 1% Toludine blue solution is used to stain suspicious looking oral mucosal lesions. Excess stain is removed after a couple of minutes using 2% acetic acid. The oral mucosa is then examined for evidence of increased staining.

Steps of Supravital staining:

Preparation of the dye:

Mashberg method of preparation of 1% Toludine blue dye:

1. One gram of Toludine blue powder

2. 10 ml of 1% acetic acid

3. 4.19 ml of absolute alcohol

4. 86 ml of distilled water

When the above ingredients are mixed it produces 1% Toludine blue solution.

The pH of the solution should be adjusted to 4.5

Preparation of oral cavity:

The patient is asked to rinse the mouth with water twice for about 20 seconds to remove debris.

1% acetic acid is given to the patient to be kept in the oral cavity for a period of 20 seconds to remove saliva which could create alkaline environment causing interpretation problems.

The toludine blue solution can be applied to the suspicious lesion with the help of cotton applicator for about 20 seconds.

Excess stain is washed off using 1% acetic acid again given as oral rinse for 20 seconds.


Dark blue staining of lesion is considered to be positive.

Light blue staining is considered rather doubtful

No staining is considered as negative


1. Staining with Toludine blue shows excellent demarcation of lesions from surrounding normal mucosa. It is hence easy to biopsy these lesions.

2. In the absence of active mucosal inflammation its sensitivity in identifying premalignant and malignant lesions is good. Its diagnostic accuracy is second only to oral cytological examination.