TY - JOUR
T1 - Early detection of apoptosis in living cells by fluorescence correlation spectroscopy
AU - Martinez, Michelle M.
AU - Reif, Randall D.
AU - Pappas, Dimitri
N1 - Funding Information:
Acknowledgements The authors would like to thank Professor Uli Nienhaus and Roger Rieger for providing us with the FCS/PCH Labview software. This work was supported in part by a grant from the Robert A. Welch Foundation (D-1667).
PY - 2010
Y1 - 2010
N2 - Early detection of apoptotic cells via caspase activity is demonstrated with fast response time. Fluorescence correlation spectroscopy (FCS) is used to identify the presence of a cleaved fluorogenic probe based on the fluorescence of rhodamine 110 in Jurkat cells. FCS curves are shown to be markedly different for autofluorescent (non-apoptotic) cells, whereas cells with cleaved probe showed diffusion and molecular brightness characteristic of rhodamine 110. Using FCS measurements, cells were identified as apoptotic on the basis of the presence of autocorrelated fluorescence, average molecular brightness (η), and molecular dwell time (τ D). Apoptotic cells identified in this manner were detected as early as 45 min after induction. Unlike other methods with similar identification times, such as western blotting and electron microscopy, cells remain viable for further analysis. This multi-parameter approach is rapid, flexible, and does not require transfection of the cells prior to analysis, enabling apoptosis to be identified early in a wide variety of cell types.
AB - Early detection of apoptotic cells via caspase activity is demonstrated with fast response time. Fluorescence correlation spectroscopy (FCS) is used to identify the presence of a cleaved fluorogenic probe based on the fluorescence of rhodamine 110 in Jurkat cells. FCS curves are shown to be markedly different for autofluorescent (non-apoptotic) cells, whereas cells with cleaved probe showed diffusion and molecular brightness characteristic of rhodamine 110. Using FCS measurements, cells were identified as apoptotic on the basis of the presence of autocorrelated fluorescence, average molecular brightness (η), and molecular dwell time (τ D). Apoptotic cells identified in this manner were detected as early as 45 min after induction. Unlike other methods with similar identification times, such as western blotting and electron microscopy, cells remain viable for further analysis. This multi-parameter approach is rapid, flexible, and does not require transfection of the cells prior to analysis, enabling apoptosis to be identified early in a wide variety of cell types.
KW - Apoptosis
KW - Cancer
KW - Caspases
KW - Fluorescence correlation spectroscopy
KW - Single-cell analysis
UR - http://www.scopus.com/inward/record.url?scp=76749093020&partnerID=8YFLogxK
U2 - 10.1007/s00216-009-3298-3
DO - 10.1007/s00216-009-3298-3
M3 - Article
C2 - 19937429
AN - SCOPUS:76749093020
VL - 396
SP - 1177
EP - 1185
JO - Analytical and Bioanalytical Chemistry
JF - Analytical and Bioanalytical Chemistry
SN - 1618-2642
IS - 3
ER -