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5x sds sample buffer become brown color while preparing
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Sudheer Sangeetham
Apr 18, 2013, 8:06:26 AM4/18/13
to Met...@magpie.bio.indiana.edu
Hello Everyone
I was trying to prepare 5x sds sample buffer and it has become brown color
instead of blue color. Could anyone please tell me what is the reason and
please tell me your suggestions.
The 5x sample buffer recipe
250mM tris-hcl ph 6.8(1M stock)------10ml
10% sds-------------------------4gr
50% glycerol(87%stock)------------------23ml
0.02% Bromphenol blue-----0.008gr
final volume-------------------------40ml with DDh20 including 1.4ml of 0.5
M B-mercaptoethanol but I didnt add here.
Thanking you in advance
--
Sudheer Babu.S
Research Fellow
Institute of Biochemistry
Biological Research Center
Szeged,Hungary.
I was trying to prepare 5x sds sample buffer and it has become brown color
instead of blue color. Could anyone please tell me what is the reason and
please tell me your suggestions.
The 5x sample buffer recipe
250mM tris-hcl ph 6.8(1M stock)------10ml
10% sds-------------------------4gr
50% glycerol(87%stock)------------------23ml
0.02% Bromphenol blue-----0.008gr
final volume-------------------------40ml with DDh20 including 1.4ml of 0.5
M B-mercaptoethanol but I didnt add here.
Thanking you in advance
--
Sudheer Babu.S
Research Fellow
Institute of Biochemistry
Biological Research Center
Szeged,Hungary.
Christian Praetorius
Apr 20, 2013, 4:36:55 PM4/20/13
to
Sudheer Sangeetham <sudhee...@gmail.com> wrote:
>I was trying to prepare 5x sds sample buffer and it has become brown color
>instead of blue color.
Two possibilities: Bromophenol blue is virtually insoluble in water.
>I was trying to prepare 5x sds sample buffer and it has become brown color
>instead of blue color.
If the concentration is too high, it stays reddish brown. Try to
dilute it with water to 1x and it should turn blue.
Then it can be a effect of the pH, since bromophenol blue is a pH
indicator as well.
Christian
--
X-no-Sig: yes
Dr Engelbert Buxbaum
Apr 21, 2013, 7:58:24 AM4/21/13
to
In article <mailman.276.136631...@net.bio.net>,
sudhee...@gmail.com says...
yellow. My guess is that your solution has a pH just around the pKa
value of bromophenolblue (3.6). Remember that the pH of most buffers
change upon dilution, because it is the activity, rather than the
concentation that determines pH. In addition, the high glycerol content
may also affect the colour. Is SDS actually soluble under these
conditions?
--
Car (noun): erratically moving obstacle on the road
sudhee...@gmail.com says...
>
> Hello Everyone
>
> I was trying to prepare 5x sds sample buffer and it has become brown color
> instead of blue color. Could anyone please tell me what is the reason and
> please tell me your suggestions.
>
> The 5x sample buffer recipe
>
> 250mM tris-hcl ph 6.8(1M stock)------10ml
> 10% sds-------------------------4gr
> 50% glycerol(87%stock)------------------23ml
> 0.02% Bromphenol blue-----0.008gr
> final volume-------------------------40ml with DDh20 including 1.4ml of 0.5
> M B-mercaptoethanol but I didnt add here.
Promophenolblue is an indicator with a colour chnage from blue to
> Hello Everyone
>
> I was trying to prepare 5x sds sample buffer and it has become brown color
> instead of blue color. Could anyone please tell me what is the reason and
> please tell me your suggestions.
>
> The 5x sample buffer recipe
>
> 250mM tris-hcl ph 6.8(1M stock)------10ml
> 10% sds-------------------------4gr
> 50% glycerol(87%stock)------------------23ml
> 0.02% Bromphenol blue-----0.008gr
> final volume-------------------------40ml with DDh20 including 1.4ml of 0.5
> M B-mercaptoethanol but I didnt add here.
yellow. My guess is that your solution has a pH just around the pKa
value of bromophenolblue (3.6). Remember that the pH of most buffers
change upon dilution, because it is the activity, rather than the
concentation that determines pH. In addition, the high glycerol content
may also affect the colour. Is SDS actually soluble under these
conditions?
--
Car (noun): erratically moving obstacle on the road
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