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Sample preparation for SDS from detergent insoluble fraction (Pellet). - (Mar/12/2011 )

Hi everyone....
i wish to screen some proteins in the detergent insolule fraction - the pellet settles after protein clarification. if i have to run this as well, how shd i prepare the sample? my plan is to add the loading dye directly and denature after mechanical disruption thru syringe, will it pull down the protein or shd i have to prepare some stronger lysis buffer inorder to see the detergent insoluble proteins in the pellet??? Kindly shed your expertise or suggestions.



if the pellet is not too large then you should be able to solubilize the proteins in sds sample buffer (with reducing agent) and heat (60-70C for 10-20 minutes or boil for ~5 minutes).


thank you....i do this and get back if i have any probs....thanks for ur reply.........