TY  - JOUR
A1  - Molesh, D A
A1  - Hall, J M
T1  - Quantitative analysis of CD34+ stem cells using RT-PCR on whole cells.
Y1  - 1994/04/01 
JF  - Genome Research 
JO  - Genome Research 
SP  - 278 
EP  - 284 
VL  - 3 
IS  - 5 
UR  - http://genome.cshlp.org/content/3/5/278.abstract 
N2  - We have employed RT-PCR of whole cells to develop a quantitative method for estimating the number of rare cells expressing a unique mRNA in a large, mixed population of cells. We have demonstrated that RT-PCR can be done on whole cells without the need for extraction of the RNA. This allows for a great saving of time and effort, as well as allowing quantitative analysis to be based on the total number of cells analyzed in a given aliquot and the presence or absence of the specific RT-PCR product. We have employed a limiting dilution series on whole cells, with multiple aliquots at each cell concentration to achieve more statistical power in the analysis of a rare cell type. We have used a nested amplification of the CD34 mRNA to be able to detect a single cell expressing the CD34 mRNA in a larger population of non-CD34-expressing cells. We demonstrate that by using this technique, cells from blood and bone marrow containing the CD34 mRNA can be followed quantitatively during a multistep purification involving immunoadsorption followed by fluorescence-activated cell sorting. We also demonstrate that many cells that express the CD34 protein on their surface no longer contain detectable levels of CD34 mRNA, a phenomenon that appears to be developmentally regulated. 
ER  -