<title>High-resolution whole field fluorescence lifetime imaging of fluorophore distribution and environment</title>

Steady-state fluorence imaging and time-resolved fluorescence imaging are two important areas in fluorescence imaging research. Fluorescence lifetime imaging is an absolute measurement method which is independent of excitation laser intensity, fluorophore concentration, and photobleaching compared to fluorescence intensity imaging techniques. Time-gated fluorescence lifetime imaging microscopy (FLIM) can provide high resolution and high imaging frame during mature FLIM methods. An abstract time-gated FLIM model was given, and important temporal parameters are shown as well. Aiming at different applications of steady and transient fluorescence processes, two different operation modes, timing and lifetime computing algorithm are designed. High resolution and high frame can be achieved by one-excitation one-sampling mode and least square algorithm for steady imaging applications. Correspondingly, one-excitation two-sampling mode and rapid lifetime determination algorithm contribute to transient fluorescence situations.

Download Full-text

High-resolution imaging of basal cell carcinoma: a comparison between multiphoton microscopy with fluorescence lifetime imaging and reflectance confocal microscopy

Skin Research and Technology ◽

10.1111/j.1600-0846.2012.00661.x ◽

2012 ◽

Vol 19 (1) ◽

pp. e433-e443 ◽

Cited By ~ 19

Author(s):

Marco Manfredini ◽

Federica Arginelli ◽

Christopher Dunsby ◽

Paul French ◽

Clifford Talbot ◽

...

Keyword(s):

High Resolution ◽

Confocal Microscopy ◽

Basal Cell Carcinoma ◽

Cell Carcinoma ◽

Fluorescence Lifetime ◽

Multiphoton Microscopy ◽

Fluorescence Lifetime Imaging ◽

High Resolution Imaging ◽

Lifetime Imaging ◽

Resolution Imaging

Download Full-text

Quantitative fluorescence lifetime imaging uncovers a novel role for KCC2 chloride transport in dendritic microdomains

10.1101/2020.06.08.139592 ◽

2020 ◽

Author(s):

Nicholas L. Weilinger ◽

Jeffrey M. LeDue ◽

Kristopher T. Kahle ◽

Brian A. MacVicar

Keyword(s):

High Resolution ◽

Fluorescence Lifetime ◽

Chloride Transport ◽

Chloride Ion ◽

Fluorescence Lifetime Imaging ◽

Lifetime Imaging ◽

Whole Cell Patch Clamp ◽

Synaptic Neurotransmission ◽

Subcellular Domains ◽

Patch Clamp Electrophysiology

AbstractIntracellular chloride ion ([Cl−]i) homeostasis is critical for synaptic neurotransmission yet variations in subcellular domains are poorly understood owing to difficulties in obtaining quantitative, high-resolution measurements of dendritic [Cl−]i. We combined whole-cell patch clamp electrophysiology with simultaneous fluorescence lifetime imaging (FLIM) of the Cl− dye MQAE to quantitatively map dendritic Cl− levels in normal or pathological conditions. FLIM-based [Cl−]i estimates were corroborated by Rubi-GABA uncaging to measured EGABA. Low baseline [Cl-]i in dendrites required Cl− efflux via the K+-Cl− cotransporter KCC2 (SLC12A5). In contrast, pathological NMDA application generated spatially heterogeneous subdomains of high [Cl−]i that created dendritic blebs, a signature of ischemic stroke. These discrete regions of high [Cl−]i were caused by reversed KCC2 transport. Therefore monitoring [Cl−]i microdomains with a new high resolution FLIM-based technique identified novel roles for KCC2-dependent chloride transport to generate dendritic microdomains with implications for disease.

Download Full-text

Abstract 15508: Random Forest Classifier-incoporated Intravascular Optical Coherence Tomography-fluorescence Lifetime Imaging (oct-flim) Provides Automated Characterization of Key Biochemical Components of Coronary Atherosclerotic Plaques

Circulation ◽

10.1161/circ.142.suppl_3.15508 ◽

2020 ◽

Vol 142 (Suppl_3) ◽

Author(s):

Sunwon Kim ◽

Hyeong Soo Nam ◽

Woo Jae Kang ◽

Joon Woo Song ◽

Hyun Jung Kim ◽

...

Keyword(s):

High Resolution ◽

Random Forest ◽

Fluorescence Lifetime ◽

Biochemical Characterization ◽

Random Forest Classifier ◽

Atherosclerotic Plaques ◽

Fluorescence Lifetime Imaging ◽

Lifetime Imaging ◽

Biochemical Components

Introduction: Fluorescence lifetime imaging (FLIm) allows label-free biochemical characterization of atheroma, however, multispectral FLIm produces massive data throughput making image interpretation problematic. We investigated whether machine learning algorithm can be applied to intravascular OCT-FLIm for automated biochemical characterization of coronary plaques. Methods and Results: We built a fully-integrated, high-speed OCT-FLIm system and a low-profile, dual-modal imaging catheter that can provide high-resolution OCT images and correctly co-registered multispectral fluorescence lifetime (FL) readouts: ch.1 and ch.2 FL, and FL intensity ratio (ch.2/ch.1). Rapid intracoronary imaging (10-20 mm/s, 100 rps) was safely performed in atheromatous pigs. There were significant differences in FL measurements according to plaque types (high-risk vs. fibrotic: p<0.001). Multispectral FL measurements sampled selectively from histologically-proven plaque components (lipid, macrophage, lipid+macrophage, SMC) were analyzed. Each component was distinguishable from one another either by difference of FLs or intensity ratio. Random forest classifier (RFC), trained with component-labeled multispectral FL dataset, accurately classified key biochemical components of atherosclerotic plaques. RFC-determined biochemical characteristics of target plaque were consistent across two repeated imaging data (intraclass correlation, p<0.0001) and corroborated closely with those derived from quantitative immunohistochemistries. Conclusions: Our OCT-FLIm incorporating RFC-based systematic multispectral FL analysis could provide high-resolution plaque imaging with automated biochemical characterization in beating coronary environment. The present imaging strategy enabling comprehensive characterization of multiple components of atherosclerotic plaques will open a new avenue in the field of cardiovascular imaging.

Download Full-text