Projects per year
Abstract
Facultative intracellular pathogens are able to live inside and outside host cells. It is highly desirable to differentiate their cellular locations for the purposes of fundamental research and clinical applications. In this work, we developed a novel analysis platform that allows users to choose two analysis models: amplitude weighted lifetime (τA) and intensity weighted lifetime (τI) for fluorescence lifetime imaging microscopy (FLIM). We applied these two models to analyse FLIM images of mouse Raw macrophage cells that were infected with bacteria Shigella Sonnei, adherent and invasive E. coli (AIEC) and Lactobacillus. The results show that the fluorescence lifetimes of bacteria depend on their cellular locations. The τA model is superior in visually differentiating bacteria that are in extra- and intra-cellular and membrane-bounded locations, whereas the τI model show excellent precision. Both models show speedy performances that analysis can be performed within 0.3 second. We also compared the proposed models with a widely used commercial software tool (τC, SPC Image, Becker & Hickl GmbH), showing similar τI and τC results. The platform also allows users to perform phasor analysis with great flexibility to pinpoint the regions of interest from lifetime images as well as phasor plots. This platform holds the disruptive potential of replacing z-stack imaging for identifying intracellular bacteria.
Original language | English |
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Article number | 034001 |
Number of pages | 10 |
Journal | Methods and Applications in Fluorescence |
Volume | 8 |
Issue number | 3 |
DOIs | |
Publication status | Published - 14 Apr 2020 |
Keywords
- fluorescence lifetime
- FLIM
- two-photon excitation
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Dive into the research topics of 'A rapid analysis platform for investigating the cellular locations of bacteria using two-photon fluorescence lifetime imaging microscopy'. Together they form a unique fingerprint.Projects
- 1 Finished
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UK Quantum Technology Hub in Quantum Enhanced Imaging (Quantic)
Birch, D., Dawson, M., Dawson, M., Strain, M., Chen, Y., Gu, E., Li, D., Strain, M., Watson, I., Jeffers, J., Oppo, G. & Yao, A.
EPSRC (Engineering and Physical Sciences Research Council), HORIBA Jobin Yvon IBH Ltd
1/10/14 → 31/10/19
Project: Research
Research output
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Emerging clinical applications of fluorescence using SPADs
Birch, D., 15 Jun 2022.Research output: Contribution to conference › Keynote
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Fluorescence guided surgery
Stewart, H. L. & Birch, D. J. S., 26 Aug 2021, In: Methods and Applications in Fluorescence . 9, 4, 26 p., 042002.Research output: Contribution to journal › Review article › peer-review
Open AccessFile16 Citations (Scopus)6 Downloads (Pure) -
TCSPC camera for real time video rate FLIM acquisition based on CMOS technology
Hungerford, G., McLoskey, D., Hirsch, R., Yip, P., Birch, D., Johnston, N. & Henderson, R., 7 Feb 2020, In: Biophysical Journal. 118, 3, Supplement 1, p. 308a-308a 1 p.Research output: Contribution to journal › Conference Contribution › peer-review
Open AccessFile2 Downloads (Pure)