Hi I would want try EM (negative stain method) with my sample but in size exclusion column I identify 3 different oligomers. Can I do EM? or I need a homogeneous sample.
Thank you
You can do EM, if you want to look at the different oligomers. If you want to determine the structure of one of the oligomers then you need to have a pure sample (recommended), or a good binning strategy to separate the 3 oligomers.
The fact that you see three different oligomers by SEC suggests very different sizes for these oligomers. Hence, you should be able to distinguish these oligomers with negative stain EM, no problem.
If the oligomers are sensitive to the buffer conditions, you might consider cross-linking before doing the negative staining, since the oligomers could fall apart during the staining. Alternatively, if you use uranyl acetate as a stain, its strong fixative properties would prevent a disassembly in most cases. Just avoid phosphate buffer in any form, since uranyl phosphate formation would ruin most staining attempts.
If you would like to study three different oligomers - you can do EM. For the specific oligomer - better to separate it.
Good luck
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