total RNA isolation from whole blood - (Aug/28/2007 )
hi,
i used fischer 344 rat whole blood in my research. i will use high pure RNA isolation kit for total RNA isolation and i must lysis red blood cell. this kit say that you use Red Blood Cell Lysis Buffer but this buffer can't lysis red blood cell in my rat's whole blood. how do i lysis rat's red blood cell?
i used fischer 344 rat whole blood in my research. i will use high pure RNA isolation kit for total RNA isolation and i must lysis red blood cell. this kit say that you use Red Blood Cell Lysis Buffer but this buffer can't lysis red blood cell in my rat's whole blood. how do i lysis rat's red blood cell?
I assume you're trying to obtain only the white blood cells. What we generally do to lyse and remove red blood cells is we had a small volume of ultra-pure water (for example 5ml) to the cell pellet. Then we mix the solution very well and add about 30ml of RPMI. After spinning the tube, a large number of the RBCs are lysed and the pellet will appear white. Often, you have to repeat this step several times.
Generally, when I'm extracting RNA from whole blood, I use the PAXgene kit from Qiagen. Unfortunately, I don't think it would be very effective for rat blood since you need to fill a whole tube with blood and buy their special equipment.
can you explain RPMI, that is RPMI expanded mean?
hi,
as far as i know means rpmi "roswell park memorial institute" and it's a name of a cell culture medium, like DMEM
How do i lysis to erytrocyte for whole blood stored -80 C but not culture medium?
and how do i determine total RNA concentration with spectrophotometer?
RPMI is a media we purchase from Quality Biological. I have no idea what it stands for, but here's a link http://www.qualitybiological.com/ptistore/...?prodId=112-040
how do i calculate total RNA concentration with spectrophotometer? i am measuring total RNA but i obtain very low absorbance??