SDS-sample buffer

Stephen C. Dahl stebby at welchlink.welch.jhu.edu
Wed Apr 15 13:00:34 EST 1998


René Bartz (bartr000 at mail.uni-mainz.de) wrote:
: Hi!

: Since severals weeks, we have a problem with our SDS-sample buffer. Our
: 1x sample buffer contains:
: 62.5 mM 0.5 M Tris Cl pH 6,8
: 10 % Glycerol
: 2% SDS
: 0.01% bromphenol blue

: The problem now is the development of flakes on the surface of the
: buffer.
: Perhaps is there an alternative protocol, which we can use.


I'll give you three  :o)

As a grad student, Len Pagliaro, another grad student at the time, gave me
the following protocol.  I'm not sure who to credit for this...

1X sample buffer

2% SDS
80 mM Tris pH 6.8
10% glycerol
.002% bromphenol blue
add 5% BME/DTT just prior to use


As a postdoc in Daniel Branton's lab we used the following protocol..

5X sample buffer

250 mM Tris pH 6.8--------------------12.5 ml 0.5M stock
10% SDS-------------------------------2.5g
50% glycerol (well, almost 50%)-------bring total volume to 25ml
.02% bromphenol blue------------------5mg
add 10% BME prior to use


A friend of mine who worked with Harvey Lodish used yet a different
protocol...

5X sample buffer

300mM Tris 6.8
25% BME
10% SDS
50% glycerol
I never asked how much BME he used

I, myself, have stuck with the Branton buffer above.  I store 500 ul
aliquots in the -20 and add 50 ul BME after thawing.  Note, however, that
this stuff is darn thick prior to the addition of the BME so I heat it at
65' C. to thaw.

As for your flakes, I can't say I've ever had that trouble.  I have been
in labs where it is so darn cold that the 10 and 20% SDS stocks come out
of solution.  These flakes tend to have neutral buoyancy though and aren't
restricted to the surface.  Maybe your SDS can't swim.

Good luck,
Steve Dahl




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