I would like to do life cell RNA-detection in T cells (human, murine, or cell lines). As SmartFlare technology from Merck relies on endocytosis, I wonder if this works in Tcells (Merck only says it might.... ). Someone with experience? Or alternative methods?
Dear Christian,
I didn't play with SmartFlare kit for RNA labelling, used RT-PCR all my life. Though ribose residues fluorescent labelling was not a fantasy 5 years ago even. But I worked with many different T-lymphocytes lines and primary cultures for 7 years (HIV cultivation) and can prove endocytosis is possible for this type of cells, even for many lines better than anywhere else. Immune response (cell or organism) is based on T-cells responsibility for alien biomaterial destroying in the body. This often happens via foregn proteins, lipids, NA and microbes captivation or "eating" - process in general very close to endocytosis. T-cells behave the same way in vitro - it is easy to feed them with any chemical or biochemical stuff.
Greetings,
E.F.
I create different cancer diagnostic kits, my impression is fluorescence-RNAs are quantitatively-reliable when it is RealTime-RT-PCR with synthetic standard calibration. The backward of the method is it is not sensitive enough for blood test diagnostic (sensitivity is high, but one cannot trust it below sertain concentration of sample). As for time lapse fluorescent antigen-specific mAbs, lipid-specific labelling, carbohydrate labelling - you should have in mind that quantification is theoretically possible but in reality difficult to achieve.
I measured pharmacocytokinetics of several antitumor agents delivery systems with confocal time lapse 10 years ago - charts and video cut-off pictures look great for publications and can be informative for decision where to move with research. Unfortunately for commercial diagnostics it is too costly or too difficult to do. So the choice of lab. kits depends on the task - not everything is even 50 percent of reality from promised in their advertisement. But for sure, the more different methods you tried to make yourself - the better is your future performance in research. And contrarywise, very often it happens that people who publish a lot do not have practical knowledge and in reality write nonsense or present negative results for positive ones. Even without intention...
Greetings
You might also be interested in this discussion on the SmarFlare technology.
https://raphazlab.wordpress.com/2015/03/11/how-smart-are-smartflares/
We have now published our article evaluating the SmartFlare technology
https://raphazlab.wordpress.com/2015/11/17/the-spherical-nucleic-acids-mrna-detection-paradox/
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