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MTT assay

MTT assay - Protocols and Methods Forum

MTT assay - Post Any Protocol, Method, Technique, Procedure or Tips / Troubleshooting for any Molecular Biology Technique.


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  #1  
Old 02-15-2006, 12:04 PM
Devendra Dusane
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Default MTT assay



Dear Dr Brookes,

I am a student from University of Pune, India, currently pursing my PhD in the field of biofilm.

I am presently working on MTT assay standardization for yeast. The assay says development of blue color by the dehydrogenase liberated by the yeast cells that cleaves MTT, in the experiment that I am conducting I am not able to get the blue color.

Can you please help me with any standard protocol for biofilm estimation with MTT or any other chemical. We also checked the MTT its fine and no problem of contamination or degradation etc is observed. We are not able to locate the problem in this assay.

We look forward for your sincere help and guidance in this.

Looking forward to hear from you soon.

Thanking you,

Sincerely

Devendra


Devendra Dusane
JRF (BARC-UoP Research Programme)
Institute of Bioinformatics and Biotechnology,
University of Pune,
Ganeshkhind, Pune: 411 007
India.
mobile: +91-9822377730
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  #2  
Old 02-20-2006, 10:51 AM
Dr Engelbert Buxbaum
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Default MTT assay

Devendra Dusane wrote:


Reference for MTT assay is Slater et al., Biochim. Biophys. Acta 77
(1963) 383. IIRC there is also a description in the methods section of
the Sigma-catalog.

MTT turns purple, not blue. Have you verified that MTT passes the yeast
cell wall? I have used it only for mammalian cells, there the method is
simple: Cells are grown in 5 mg/ml MTT solution in medium for 3-4 h, the
medium is discarded, cells briefly washed with PBS and the pigment
dissolved in 50 mM HCl in i-propanol. Photometric measurement is done at
570 nm.
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