Methyl Red Test Principle,Procedure and Result interpretation

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Methyl red test MLTGEEKS

MR Test is a quantitative biochemical tests for identification of bacteria by detection of acid production which requires positive organisms to produce strong acid (lactic, formic) from glucose through the mixed acid fermentation pathway.Methyl red is a pH indicator having a range between 6.0(yellow) and 4.4(red). It is used in IMViC test for detection of acid production from glucose fermentation. Since many species of the family of Enterobacteriaceae may produce sufficient quantities of strong acid that can be detected by methyl red indicator during the initial phases of incubation,only those organisms that are able to maintain this low pH after prolonged incubation (48-72 hours) and overcoming the pH buffering system of the medium can be called as methyl red positive.

Principle of Methyl red Test

  • The development of stable red colour in surface of the medium indicates sufficient acid production which lower the  pH to 4.4 and constitutes positive test.
  • MR negative organisms may produce lesser quantities of acid from test substrate, yielding an intermediate orange colour between yellow and red may developed

Composition of Methyl red Test

Methyl red0.200 gm
Distilled water40.000 ml
Ethyl alcohol60.000 ml

Appearance of MR indicator

It has a pinkish red coloured solution.

Methyl Red Indicator
Methyl Red Indicator

Methyl red Test Procedure

  • Start by adding 1-2 drops of methyl red indicator  to a 5 ml of a 24 – 48 hours old culture of the suspected organism
  • Observe and interpret the colour change immediately.

Result and interpretation

Interpretation of  the results is done immediately after the addition of Methyl red indicator

Positive Methyl Red Organismred colouration to the medium
Negative Methyl Red Organismyellow colour
Also read  Phenylalanine Deamination test procedure and result interpretation
Left Methy Red positive ( Escherichia coli)-Right Methy Red Negative (Enterobacter cloacea)
Left Methy Red positive ( Escherichia coli) – Right Methy Red Negative (Enterobacter cloacea)

Culture observation and Methyl Red reaction

These cultural responses is observed after  18-24 hours incubation at temperature 37°C in MR-VP medium followed by Methyl Red test performed by using Methyl red indicator.

OrganismMethyl Red Reaction
Klebsiella pneumoniaeNegative (Yellow colour observed)
Enterobacter aerogenesNegative (Yellow colour observed)
Escherichia coliPositive (Red colour observed)


  • Color Atlas and Textbook of Diagnostic Microbiology,4th edition ,Elmer W. Koneman. Stephen D.Allen. ,William M. Janda., Paul C. Schreckenberger.,Washington C.Winn.

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About the Author: Arthur Westmann

DEFFE ARTHUR (AMOEBAMANN) is the founder and author of MLTGEEKS and MLTEXPO.He’s from Cameroon and is currently a Final year State Medical Laboratory Technician (MLT MA). Beyond lab works, he’s a passionate internet user with a keen interest in web design and blogging. Furthermore He likes traveling, hanging around with friends and social networking to do in his spare time.

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