2Mercapto ethanol protocol

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zian's picture
2Mercapto ethanol protocol

Hi sir...I Will be thank-full if you answer of my question
my question is can I use 2ME (stock) with its dilution that used in PCR protocol  for example(50microliter2me stock diluted in 5 ml diluent) for destruction of IgM  Ab in serum for toxoplasmosis.If yes, how much time  incubation. If not  please tell me the correct concentration for dilution of stock2me with incubation period with temperature?  with my regard

protoldo's picture
Hola, as you know if you add

Hola, as you know if you add 2ME to a serum, you will reduce all the proteins and not only IgM. The amount to be add will depends of the concentration of the sample. With a pure protein, Knowing the concentration and the number of S-S bridges you could estimate the amount to add for a total or partial reduction. Well, the commercial reagent is about 15 Molar, a 1/1000 dilution (1ul/1ml) will be 15 mM and the reduction is the moment of addition without the need of boiling. Concentrations about 10mM are used for routine purification steps. about,50mM are used in processes of purification under denaturant conditions (with urea or guanidine HCl) and 2%  (300mM) and boiling is used for total denaturation for PAGE. Good luck