Time-domain Fluorescence Lifetime Imaging Microscopy (FLIM) is a remarkable tool to monitor the dynamics of fluorophore-tagged protein domains inside living cells. We propose a Wide-Field Multi-Parameter FLIM method (WFMP-FLIM) aimed to monitor continuously living cells under minimum light intensity at a given illumination energy dose. A powerful data analysis technique applied to the WFMP-FLIM data sets allows to optimize the estimation accuracy of physical parameters at very low fluorescence signal levels approaching the lower bound theoretical limit. We demonstrate the efficiency of WFMP-FLIM by presenting two independent and relevant long-term experiments in cell biology: 1) FRET analysis of simultaneously recorded donor and acceptor fluorescence in living HeLa cells and 2) tracking of mitochondrial transport combined with fluorescence lifetime analysis in neuronal processes.

Download full-text PDF

Source
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3032730PMC
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0015820PLOS

Publication Analysis

Top Keywords

living cells
12
wide-field multi-parameter
8
multi-parameter flim
8
fluorescence lifetime
8
flim long-term
4
long-term minimal
4
minimal invasive
4
invasive observation
4
observation proteins
4
living
4

Similar Publications

Want AI Summaries of new PubMed Abstracts delivered to your In-box?

Enter search terms and have AI summaries delivered each week - change queries or unsubscribe any time!