Dear coordinates
i need a little help. i have determined the concentration of protein and then added equal volume of loading buffer 2X. i want to make sure that loading buffer will dilute the concentration of proteins within the sample or not? if i want to collect 30ug from that sample (3ug/ul) than i have to pick 10ul of sample or 20ul. its initial concentration before addition of loading buffer was 3ug/ul. i am little bit confused with addition of loading buffer.
waiting for your kind reply.
Regards
I am not quite sure about the second half of your question.
However, if you add an equal volume of 2x buffer to your sample, then the sample has afterwards 1/2 the concentration and the buffer is 1x.
Mohsin Nawaz
With any concentration calculation you can use the following:
(Initial Concentration) x (Volume to be Added) = (Final Concentration) x (Final Volume) or M1V1=M2V2. And use some algebra to solve for required volume of initial concentration to be added. But if you are only using a 2x solution, 1:1 ration of sample and buffer will bring it to 1x. So for example. Your protein concentration is 3ug/ul (M1). You want the final volume to be 20 uL (V2) and Final Concentration as 1.5 ug/uL (M2). You solve for V1, which would result in 10uL required of your protein solution. This is very useful when you are not doing just a 1/2 dilution.
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