Autofluorescence lifetime

A typical result is shown in Fig. 5.67. It shows autofluorescence lifetime images of stratum comeum (upper row, 5 pm deep), and stratum spinosum (lower row, 50 mm deep). The multiexponential decay was approximated by a doubleexponential model, and the decay parameters determined by a Levenberg-Marquardt fit. The colour represents the fast lifetime component, Ti, the slow lifetime component, i2, the ratio of lifetime components, Zi /12, and the ratio of the amplitudes of the components, fli / fl2- The brightness of the pixels represents the intensity. 

Fig. 5.78 Two-photon autofluorescence lifetime image of aortic tissue. Top Intensity image, lifetime image of average lifetime, lifetime distribution. Bottom Fluorescence decay in indicated pixel and double exponential fit with 81% of 294 ns and 18.7% of 2.26 ns. From [39]...

D. Schweitzer, A. Kolb, M. Hammer, Autofluorescence lifetime measurement in images of the human ocular fundus, SPIE 4432, 29-39 (2001)... 

It has been found that tumors can be detected in a very early stage of development by pretreatment with porphyrin photosensitizers. The tumor-cells display a marked increased fluorescence lifetime as compared to surrounding control cells [151-154]. Also lifetime-differences in normal autofluorescence have been detected between normal and tumor-cells. Recent studies revealed that FLIM could assist in histopathological assessment of fixed (unstained) breast cancer tissues. The autofluorescence lifetime contrast revealed statistically relevant differences (p<0.05) between benign and mahgnant stroma samples [155]. 

Raluca Niesner, B. P., Schliische, P. and Gericke, K. H. (2004). Noniterative Biexponential fluorescence lifetime imaging in the investigation of cellular metabolism by means of NAD(P)H autofluorescence. Chem. Phys. Chem. J, 1141-9. 

How can phase-modulation fluorometry contribute to this health-care need It now seems possible to construct a lifetime-based blood gas catheter (Figure 1.3), or alternatively, an apparatus to read the blood gas in the freshly drawn blood at the patient s bedside. To be specific, fluorophores are presently known to accomplish the task using a 543-nm Green Helium-Neon laser,(18 19) and it seems likely that the chemistries will be identified for a laser diode source. The use of longer wavelengths should minimize the problems of light absorption and autofluorescence of the samples, and the use of phase or modulation sensing should provide the robustness needed in a clinical environment. For the more technically oriented researcher, we note that the... 

In addition, the luminescence lifetime of quantum dots is approximately one order of magnitude longer than that of their organic counterparts.30 Thus, the autofluorescence of biological samples can effectively be filtered, for example, with time-gated measurements, in the case of these inorganic luminophores. Moreover, QDs are also particularly interesting for multiplexed analysis because of two of their inherent... 

The same publication indicated that fluorescence lifetimes of compounds from the compound collection of Pfizer classified as problematic due to their autofluorescence characteristics resulted in false positive results in many FI-based assays. For most compounds, the fluorescence lifetimes were below 1 ns. Thus, FLT measurements with a reporter fluorophore displaying a lifetime significantly longer than 1 ns are suitable for application in protease assays for compound testing. However, for applications under initial velocity conditions with a substrate turnover below 20%, fluorophores with lifetimes of a few nanoseconds are still critical because the dynamic range of the assay is then too low with lifetimes below 1 ns. 

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