The squared distance approach to frequency domain time‐resolved fluorescence analysis

Abstract

A frequency‐domain (FD) analysis of fluorescence lifetime (FLT) is a unique and rapid method for cellular and intracellular classifications that can serve for medical diagnostics purposes. Nevertheless, its data analysis process demands nonlinear fitting algorithms that may distort the resolution of the FLT data and hence diminish the classification ability of the method. This research suggests a sample classification technique that is unaffected by the analysis process as it is based on the squared distance (D2) between the raw frequency response data (FRD). In addition, it presents the theory behind this technique and its validation in two simulated data sets of six groups with similar widely and closely spaced FLT data as well as in experimental data of 43 samples from bacterial and viral infected and non‐infected patients. In the two simulated tests, the classification accuracy was above 95% for all six groups. In the experimental data, the classification of 41 out of 43 samples matched earlier report and 29 out of 31 agreed with preliminary physician diagnosis. The D2 approach has the potential to promote FD‐time resolved fluorescence measurements as a medical diagnostic technique with high specifity and high sensitivity for many of today's conventional diagnostic procedures.

Real‐time fiber‐based fluorescence lifetime imaging with synchronous external illumination: A new path for clinical translation

“…Overall, these results highlight the benefit of the phasor approach for our implementation: phasors can provide robust and online feedback of the fluorescence decay measurements in an implementation with such strict timing requirements as ours; it provides means for rapid and online‐compatible segmentation within the phasor map to highlight specific regions within the fluorescence image, as demonstrated in Figures H and S12. We note that other fluorescence lifetime data analysis methods such as multi‐exponential fitting , rapid lifetime determination , Laguerre expansion or the D 2 approach could also be employed, provided that they can meet the timing requirements of this implementation. In the future, we aim to investigate the suitability of these methods to provide real‐time feedback in our imaging platform.…”

Imaging the rotational mobility by frequency domain time-resolved fluorescence anisotropy

“…Therefore, FLIM is popular for investigating biochemical reactions and molecular interactions within the cellular environment that are valuable for biological and medical imaging [10,11]. FLIM was used for the identification of chromosomal abnormalities in Leukemias [12], or carriers of BRCA mutations [13] and detection of metastatic cells in the cerebrospinal fluid of children with Medulloblastoma [14] or the presence of viral and bacterial pathogens [15] using the crossing point method [16] and the D 2 technique [17].…”

Probing Polarity and pH Sensitivity of Carbon Dots in Escherichia coli through Time-Resolved Fluorescence Analyses

“…Previously, we demonstrated a fast method using FD fluorescence lifetime (FLT) imaging microscopy (FLIM) for detecting bacterial and viral pathogens, including E. coli [ 37 , 38 ]. FLT offers a distinctive approach to investigating complex biological environments and uncovering molecular heterogeneity by tracking differences in the excited state kinetics of one or more fluorophores [ 27 ].…”