# lysis buffer preparation - buffer preparation (Aug/01/2007 )

Hi
I want to make lysis buffer in 1 litre

400 mM Nacl
0.5% tritonX-100
50 mM tris ph 7.4

can anyone tell me how to do it?

-julee-

You need to know the concentrations of your stock solutions (NaCl and Tris) before calculating the exact amount of each needed to achieve the final concentrations.

For now, the only thing you can calculate is your 0.5% Triton, since it is considered to be 100% stock solution.

It is not mentionned, but the 0.5% is volume/volume. So you just calculate 0,5% of 1000 ml, wich gives you 5ml.

for1liter, weight 23,37 grams of NaCl (58,44*0,4) in a becher
with magnetic stirrer homogenize till dissolution.
I assume you've prepared a 1M Tris HCl pH7.4 solution.
add water up to roughly 900ml
add 5ml triton X100 (a 100%solution).
transfer all in a gratuated "burette" (can't find the english word so i've attached the picture.
Rinse becher with water and put the rinsing water in the burette. redo till getting 1L

sterilize either by filtering or autoclave (cheaper)

-fred_33-

thanks a lot

QUOTE (fred_33 @ Aug 1 2007, 06:59 AM)
for1liter, weight 23,37 grams of NaCl (58,44*0,4) in a becher
with magnetic stirrer homogenize till dissolution.
I assume you've prepared a 1M Tris HCl pH7.4 solution.
add water up to roughly 900ml
add 5ml triton X100 (a 100%solution).
transfer all in a gratuated "burette" (can't find the english word so i've attached the picture.
Rinse becher with water and put the rinsing water in the burette. redo till getting 1L

sterilize either by filtering or autoclave (cheaper)

-julee-

hi
for nacl, it says 99% and for tris it says 99.9%
the m.w. of nacl is 58.4 and that of tris is 121.1
will that help?

QUOTE (Madrius @ Aug 1 2007, 06:11 AM)
You need to know the concentrations of your stock solutions (NaCl and Tris) before calculating the exact amount of each needed to achieve the final concentrations.

For now, the only thing you can calculate is your 0.5% Triton, since it is considered to be 100% stock solution.

It is not mentionned, but the 0.5% is volume/volume. So you just calculate 0,5% of 1000 ml, wich gives you 5ml.

-julee-

fred 33 has already given you the amount of NaCl to weigh in order to have 400mM.

As for the Tris, you will have to prepare a solution of 1M and adjust the pH at 7.4. Then, as fred 33 mentionned, you have to take 50 ml of that solution and add it to the water, add the 23,37 grams of NaCl, the 5 ml of Triton and finally adjust to 1000ml.

Hope this helps,