I tried precipitation with cold acetone several times but it didn't work. I tried to use dialysis for several days but my H-NMR analysis shows traces of NHS. How can I get rid of the reagents which did not participate in the reaction?
What did you try to precipitate? Please describe your experiment in more detail. EDC and NHS are highly water and acetone soluble and should be no problem to be removed.
hey Michael,
I was trying to precipitate the product of my reaction based on few methods recommended by articles. They say that by precipitation using cold acetone we can extract the product in the form of sediment.
If your product is a protein conjugate or other polymer, you may remove EDC and NHS either by dialysis or preferably by size exclusion chromatography, e.g. by PD-10 desalting columns or equivalent.
I even tried desalting columns for removal of EDC and NHS but it is not completely removed. I had tried COOH activation on DNA but no results. has anyone activated COOH on DNA with EDC and NHS?
Most scientists do not use desalting colums optimally, because the protocols are somehow misleading. In these cases, some impurities remain. How did you perform your desalting step?
I've used gel filtration column. Basically, my aim was buffer exchange.
If NHS and EDC is still present, you most likely did not perform it properly.
I have used 1mL column. Added 0.5mL dd. water every time to wash the column. After washing step for about 1 hour, I used to wait for all dd. water to get eliminated upto the gel mark and then slowly added sample. Next, added 0.5mL distilled water, waited for water to come to the mark of gel, added 0.5mL distilled water again and this time collected my sample. This is the way i performed by gel filtration step. If there is any other way, kindly suggest.
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