Hello,everyone,
I’m a graduate student.I used the worm to study the aging.Now,I am in trouble for protein quantitation of the nematode.Does anyone to know how to do it?
Any tips or suggestions will be very helpful for me. Thank you!
We use a pretty simple method of quantifying worm protein from extracts. We grind worms in liquid nitrogen, spin out debris, and then spot aliquots of 1-5 ul onto a wetted piece of PVDF membrane (methanol/acetic acid) alongside of aliquots of BSA from 0.25 to 5 ug (in the same volume as your extract aliquot). Let the membrane dry completely, then stain with Coomassie brilliant blue (10 min) and destain several times briefly. The estimates of spot intensity are usually more accurate for us than a solution Bradford assay, and are not as easily disrupted by other reagents because the protein sticks to the blot while soluble chemicals are washed away.
bdk