:: Search forums :: Top Users :: Reagent
Position: Discussion forums > Bioscience biotechniques discussion forums > Proteomics and protein biochemistry forum
|
Nuclear Fraction Marker
Posted by: Larus50sce (IP Hidden, New member, 2)
Date: April 28, 2005 02:54PM
I am trying to fractionate SH-SY5Y cells (human neuroblastoma cell line) into
cytosol and soluble nuclear fractions, and I need to find a protein for which I could immunoblot to check that I didn't contaminate the fractions with each other. I use tubulin as a cytoplasm marker, but I am having trouble coming up with a nuclear fraction marker - something specifically localized to the nucleus, and absent in the cytoplasm. Histones would be the obvious choice, but I am lysing the cells by swelling on ice in different amounts of Triton-X-100 (0.1% for cytosol, 1% for the nuclei) because I need the lysate for immunoprecipitation and RT-PCR, so my lysis conditions may be too mild to release histones into the soluble nuclear fraction. Does anybody have a suggestion for a good nuclear fraction marker? What do other people use in such circumstances? Thanks a lot.
Re: Nuclear Fraction Marker
Posted by: 5'-ATCG (IP Hidden, Unregistered user, )
Date: April 28, 2005 07:24PM
Another hallmark is laminin B.
Re: Nuclear Fraction Marker
Posted by: Larus50sce (IP Hidden, New member, 2)
Date: April 29, 2005 12:22PM
Laminin is an extacellular matrix protein, as far as I know. How is it a marker of the nuclear fraction? Is it present in the nuclear matrix also?
Re: Nuclear Fraction Marker
Posted by: 5'-ATCG (IP Hidden, Unregistered user, )
Date: April 29, 2005 04:05PM
Please check the literatures......."laminin-B" in the nuclear lamina.
Don't get confused with laminin-1 or laminin-domain or laminin repeats...... Am J Physiol Gastrointest Liver Physiol 1999, 277: G1207-G1216; Journal of Cellular Physiology 2003, 197: 181 - 188; Neuropathology & Applied Neurobiology 2004, 30: 315... and many many more..................................................
Re: Nuclear Fraction Marker
Posted by: aniu1229 (IP Hidden, New member, 8)
Date: July 11, 2005 11:12AM
I noticed that there are lamin A, B and C, many kinds of antibody available in santacruz. do you know what is the difference among them?
Because I saw in the paper, someone used Lamin B as nuclear marker, while someone used laminA/C. if you go through the catalog of santacruz, you will find that some lamin localized in nuclear envelop, while some localized all over the nucleus. so do u think which one is better to be used as a marker? whether proteins localizing to the nuclear envelop will be easy to go to cytoplasm?
|
We are moving ... Please post to our new community forums
