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Sample buffer

sample buffer DTT Western Blot Immunoblotting

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#1 Mad Researcher

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Posted 05 February 2018 - 04:31 PM

Hi,

 

I have 800 µl of sample buffer stocks (without DTT) stored at -20°C. Before using I add 2.5 M DTT and use it.

 

Sometimes, I add the sample buffer to my samples and store at -20°C. However, the next day after heating the samples, the color of the buffer changes from blue to dark green. But, when I load it to the gels, they appear blue. Why does the color change after heating. 

 

After adding the buffer to the sample, should I store the buffer at -20°C for next day use or should I store at RT. How long can the buffer be stored (days, weeks?) at RT or -20°C?

 

Thanks.


Edited by Mad Researcher, 05 February 2018 - 07:25 PM.

Cheers,

Mad Researcher

#2 bob1

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Excellent

Posted 07 February 2018 - 08:47 AM

The colour change indicates a pH change. The dye is yellow at pH 3 and blue at pH 4.6, so it indicates that you have an acidic solution, pushing the pH lower, probably somewhere around pH 4. 

 

The buffer is best stored frozen at -20 and can be freeze/thawed a few times. If it starts to do the pH change thing - discard it.







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