Jump to content

  • Log in with Facebook Log in with Twitter Log in with Windows Live Log In with Google      Sign In   
  • Create Account

Submit your paper to J Biol Methods today!
Photo
- - - - -

Yellow Laemmli sample buffer


  • Please log in to reply
6 replies to this topic

#1 agriman

agriman

    member

  • Members
  • Pip
  • 4 posts
0
Neutral

Posted 07 July 2009 - 09:03 PM

Dear all,

I prepared 5X Laemmli sample buffer. But the color is yellow which means low pH. When I mix my sample with loading buffer, the color is still yellow.

Mixture of Sample buffer is:

2M Tris-HCl (pH 6.8) 125 ul
SDS 100 mg
Beta-Mercapt. 300 ul
Glycine 500 ul
BPB (1%) 20 ul
DW 55 ul
Total 1000 ul


I am beginner in this field. Any help and advice would be great...

#2 Minnie Mouse

Minnie Mouse

    Super Mouse

  • Global Moderators
  • PipPipPipPipPip
  • 79 posts
10
Good

Posted 07 July 2009 - 10:26 PM

You may wish to check the pH of the 2M Tris-HCl stock solution.

Hope this may help.

#3 agriman

agriman

    member

  • Members
  • Pip
  • 4 posts
0
Neutral

Posted 08 July 2009 - 02:46 AM

Minnie Mouse,

Thank you for replying to my post.

Yes, I just checked the pH of 2M Tris-HCl again and it was around 6.86. But, after adding SDS, pH is decrease. Do you think, I should adjust the pH at 6.8 after adding SDS?

Thanks...

#4 mdfenko

mdfenko

    an elder

  • Active Members
  • PipPipPipPipPipPipPipPipPipPip
  • 2,799 posts
133
Excellent

Posted 08 July 2009 - 09:44 AM

Minnie Mouse,

Thank you for replying to my post.

Yes, I just checked the pH of 2M Tris-HCl again and it was around 6.86. But, after adding SDS, pH is decrease. Do you think, I should adjust the pH at 6.8 after adding SDS?

Thanks...


i think you should replace the sds.

in your recipe you say you add 500ul glycine, did you mean glycerol? if not then that could account for the low pH.

Edited by mdfenko, 08 July 2009 - 09:46 AM.

talent does what it can
genius does what it must
i do what i get paid to do

#5 agriman

agriman

    member

  • Members
  • Pip
  • 4 posts
0
Neutral

Posted 09 July 2009 - 09:41 PM

I replaced the SDS with new one, then I prepared new sample buffer, new running buffer and new gel. All problems were disappeared. No yellow color in sample buffer, no smiling effect, no heating problem etc.

Thank you four your advice...

#6 Kriz

Kriz

    member

  • Active Members
  • Pip
  • 5 posts
0
Neutral

Posted 31 August 2010 - 06:51 AM

I replaced the SDS with new one, then I prepared new sample buffer, new running buffer and new gel. All problems were disappeared. No yellow color in sample buffer, no smiling effect, no heating problem etc.

Thank you four your advice...




I'm also new in this field. I prepared sds reducing sample buffer and the colour of the buffer was yellow. Irepeated the preparation and again it was yellow but as soon as I added it to my protein lysate, it turned blue. Is this correct?

#7 mdfenko

mdfenko

    an elder

  • Active Members
  • PipPipPipPipPipPipPipPipPipPip
  • 2,799 posts
133
Excellent

Posted 31 August 2010 - 09:36 PM

does it turn yellow after the addition of sds? if so then your sds is old and decomposing.

if it's yellow before adding sds then your buffer is not made correctly (tris).
talent does what it can
genius does what it must
i do what i get paid to do




Home - About - Terms of Service - Privacy - Contact Us

©1999-2013 Protocol Online, All rights reserved.