hi im a noobie when it comes to lab work but recently i made a careless mistake and i dont think i can afford to throw it all away and make a new set of buffer due to lack of starting ingredients.
firstly, i was supposed to prepare the 10x nitrocellulose transfer buffer for western blot in the following recipe,
10X Nitrocellulose transfer buffer:
· Tris base 30.3 g
· Glycine 144g
· miliQ 1L
However, i only added 800ml of miliQ unknowingly forgotten to top it up to 1L of miliQ after dissolving.
I then proceeded to prepare the 1X nitrocelluose transfer buffer in the following recipe,
1X Nitrocellulose transfer buffer:
· 200 ml '10X' nitrocellulose transfer buffer (not really 10X nitrocellulose transfer buffer, probably more)
· 200 ml methanol
· 1.6L miliQ
would my 1X nitrocellulose transfer buffer work?
Hi,
Your transfert buffer will work even if it is more concentrate. Maybe the transfert will be less efficient.
In fact, you have 12.5X transfer buffer, if you can, you just need to reajust the 1X solution by adding 50mL methanol and 450mL miliQ (final solution 2,5 L, 10X methanol = 250 mL, 12.5X Buffer = 200 mL, 2.05L miliQ).
The short answer is YES. However, if you are interested in transfer of small molecular weight proteins (below 50 kDa), decrease the transfer time slightly.
Yes !
For 200mL of a 12.5X solution buffer, you need 2500mL for 1X final (2500/12.5 = 200). So just ajust your last solution up to 2500mL, add 50mL methanol (10X ) and 450 mL miliQ to complete .
But, as we say with Divader, your first solution will work anywy.
Regards
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