PMID- 14706924
OWN - NLM
STAT- MEDLINE
DCOM- 20040810
LR  - 20080815
IS  - 1121-760X (Print)
IS  - 1121-760X (Linking)
VI  - 47
IP  - 4
DP  - 2003
TI  - Dual excitation multi- fluorescence flow cytometry for detailed analyses of 
      viability and apoptotic cell transition.
PG  - 289-98
AB  - The discrimination of live/dead cells as well as the detection of apoptosis is a 
      frequent need in many areas of experimental biology. Cell proliferation is linked 
      to apoptosis and controlled by several genes. During the cell life, specific 
      events can stimulate proliferation while others may trigger the apoptotic 
      pathway. Very few methods (i.e. TUNEL) are now available for studies aimed at 
      correlation between apoptosis and proliferation. Therefore, there is interest in 
      developing new methodological approaches that are able to correlate apoptosis to 
      the cell cycle phases. Recently new approaches have been proposed to detect and 
      enumerate apoptotic cells by flow cytometry. Among these, the most established 
      and applied are those based on the cell membrane modifications induced in the 
      early phases of the apoptotic process. The dye pair Hoechst 33342 (HO) and 
      Propidium Iodide (PI), thanks to their peculiar characteristics to be 
      respectively permeable and impermeable to the intact cell membrane, seems to be 
      very useful. Unfortunately the spectral interaction of these dyes generates a 
      consistent "energy transfer" from HO to PI. The co-presence of the dyes in a 
      nucleus results in a modification in the intensity of both the emitted 
      fluorescences. In order to designate the damaged cells (red fluorescence) to the 
      specific cell cycle phases (blue fluorescence), we have tested different staining 
      protocols aimed to minimize the interference of these dyes as much as possible. 
      In cell culture models, we are able to detect serum-starved apoptotic cells as 
      well as to designate their exact location in the cell cycle phases using a very 
      low PI concentration. Using a Partec PAS flow cytometer equipped with HBO lamp 
      and argon ion laser, a double UV/blue excitation has been performed. This 
      analytical approach is able to discriminate live blue cells from the damaged 
      (blue-red) ones even at 0.05 micro g/mL PI. The same instrumental setting allows 
      performing other multi-colour analyses including AnnexinV-FITC as well as the 
      possibility to make a correlated analysis to phenotype markers.
FAU - Mazzini, G
AU  - Mazzini G
AD  - Istituto di Genetica Molecolare del CNR, Sezione di Istochimica e Citometria, 
      Piazza Botta 10, 27100 Pavia, Italy. mazzi@igm.cnr.it
FAU - Ferrari, C
AU  - Ferrari C
FAU - Erba, E
AU  - Erba E
LA  - eng
PT  - Journal Article
PL  - Italy
TA  - Eur J Histochem
JT  - European journal of histochemistry : EJH
JID - 9207930
RN  - 0 (DNA, Neoplasm)
SB  - IM
MH  - *Apoptosis
MH  - Cell Line, Tumor
MH  - *Cell Survival
MH  - DNA, Neoplasm/analysis
MH  - Flow Cytometry/instrumentation/*methods
MH  - Humans
MH  - Leukemia, T-Cell/*pathology
MH  - Microscopy, Fluorescence/instrumentation/*methods
MH  - Ploidies
EDAT- 2004/01/07 05:00
MHDA- 2004/08/11 05:00
CRDT- 2004/01/07 05:00
PHST- 2004/01/07 05:00 [pubmed]
PHST- 2004/08/11 05:00 [medline]
PHST- 2004/01/07 05:00 [entrez]
PST - ppublish
SO  - Eur J Histochem. 2003;47(4):289-98.