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146-EN, General section

Use of methylene blue as staining agent to reveal DNA


Methylene blue may be used as an alternative to ethidium bromide, a suspected mutagen,  in staining  Agarose gel  to reveal DNA.

Substituted substance(s)

  1. Ethidium bromide

    CAS No. 1239-45-8 EC No. 214-984-6 Index No. 612-278-00-6

Alternative substance(s)

  1. Methylene blue

    CAS No. 61-73-4 EC No. 200-515-2 Index No. 

Other type of alternative

Reliability of information

Internet information: data are from an internet document and only a basic and partial evaluation could be performed

Hazard assessment

Methylene (methylthioninium chloride) blue is not classified as hazardous according to the harmonized classification nor is it notified in the C&L inventory. See also Further information.

» Check the Substance Database according to SUBSPORT Screening Criteria (SDSC)

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Substitution description

Electrophoresis of DNA requires a staining step to make the separated DNA fragments visible. The
safest and easiest method is to use methylene blue solution. As methylene blue carries a positive
charge, it binds to the negatively charged DNA fragments and stains them blue. Two methods may be used: with diluted or with concentrated methylene blue. The method using diluted methylene blue involves submerging the gel in   methylene blue solution (0.025%)   and allowing it to    stand for up to 16 hours so the stain diffuses into the gel and  binds with the DNA fragments.

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Case/substitution evaluation

Methylene blue is a commonly used and available chemical, whose use in revealing DNA does not involve high costs or complicated laboratory procedures.It is a good substitution especially for school laboratories.

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Other solutions

Further information

 MSDS  methylene blue (methylthioninium chloride) from Sigma -Aldrich

Further languages available

Who provided the information

Type of information supplier
Producer / distributor


Southern Biological, a division of Cogitamus Pty Ltd, Southern Biological
19-21 Worrell Street, Nunawading,Victoria 3131, Australia,

Publication source

Southern Biological, Australia.

Type of publication and availability 

Document freely available on internet

Original document: click here
Archived copy: click here

Comments from SUBSPORT users

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