Laser microscopy of

1998 ◽  
Author(s):  
Kazuto Ikeda ◽  
Takahiro Kimura ◽  
Tomoji Nakamura
Keyword(s):  
Laser Microscopy

scholarly journals Femtosecond pulsed laser microscopy: a new tool to assess the in vitro delivered dose of carbon nanotubes in cell culture experiments

2021 ◽  
Vol 18 (1) ◽  
Author(s):  
Dominique Lison ◽  
Saloua Ibouraadaten ◽  
Sybille van den Brule ◽  
Milica Todea ◽  
Adriana Vulpoi ◽  
...  
Keyword(s):  
Carbon Nanotubes ◽  
Cell Culture ◽  
Pulsed Laser ◽  
Lung Fibroblasts ◽  
Laser Microscopy ◽  
Cell Responses ◽  
The Impact ◽  
Delivered Dose ◽  
Femtosecond Pulsed Laser

Abstract Background In vitro models are widely used in nanotoxicology. In these assays, a careful documentation of the fraction of nanomaterials that reaches the cells, i.e. the in vitro delivered dose, is a critical element for the interpretation of the data. The in vitro delivered dose can be measured by quantifying the amount of material in contact with the cells, or can be estimated by applying particokinetic models. For carbon nanotubes (CNTs), the determination of the in vitro delivered dose is not evident because their quantification in biological matrices is difficult, and particokinetic models are not adapted to high aspect ratio materials. Here, we applied a rapid and direct approach, based on femtosecond pulsed laser microscopy (FPLM), to assess the in vitro delivered dose of multi-walled CNTs (MWCNTs). Methods and results We incubated mouse lung fibroblasts (MLg) and differentiated human monocytic cells (THP-1) in 96-well plates for 24 h with a set of different MWCNTs. The cytotoxic response to the MWCNTs was evaluated using the WST-1 assay in both cell lines, and the pro-inflammatory response was determined by measuring the release of IL-1β by THP-1 cells. Contrasting cell responses were observed across the MWCNTs. The sedimentation rate of the different MWCNTs was assessed by monitoring turbidity decay with time in cell culture medium. These turbidity measurements revealed some differences among the MWCNT samples which, however, did not parallel the contrasting cell responses. FPLM measurements in cell culture wells revealed that the in vitro delivered MWCNT dose did not parallel sedimentation data, and suggested that cultured cells contributed to set up the delivered dose. The FPLM data allowed, for each MWCNT sample, an adjustment of the measured cytotoxicity and IL-1β responses to the delivered doses. This adjusted in vitro activity led to another toxicity ranking of the MWCNT samples as compared to the unadjusted activities. In macrophages, this adjusted ranking was consistent with existing knowledge on the impact of surface MWCNT functionalization on cytotoxicity, and might better reflect the intrinsic activity of the MWCNT samples. Conclusion The present study further highlights the need to estimate the in vitro delivered dose in cell culture experiments with nanomaterials. The FPLM measurement of the in vitro delivered dose of MWCNTs can enrich experimental results, and may refine our understanding of their interactions with cells.

125 CONFOCAL LASER MICROSCOPY FOR DIAGNOSIS OF UROTHELIAL BLADDER CANCER: A TECHNOLOGY OF THE FUTURE?

2009 ◽  
Vol 8 (4) ◽  
pp. 152
Author(s):  
C. Wiesner ◽  
A. Salzer ◽  
R. Kiesslich ◽  
C. Hampel ◽  
R. Gillitzer ◽  
...  
Keyword(s):  
Bladder Cancer ◽  
Confocal Laser Microscopy ◽  
Confocal Laser ◽  
Urothelial Bladder Cancer ◽  
Laser Microscopy ◽  
The Future ◽  
Urothelial Bladder

Laser Microscopy of Scattering Media Based on the Regularized Minimally Diffuse Image Reconstruction

JETP Letters ◽  
2021 ◽  
Vol 114 (8) ◽  
pp. 451-455
Author(s):  
M. S. Pochechuev ◽  
I. V. Fedotov ◽  
E. A. Stepanov ◽  
M. S. Andreeva ◽  
A. B. Fedotov ◽  
...  
Keyword(s):  
Image Reconstruction ◽  
Scattering Media ◽  
Laser Microscopy

scholarly journals Assessment of Fluorochromes for Two-Photon Laser Scanning Microscopy of Biofilms

2002 ◽  
Vol 68 (2) ◽  
pp. 901-909 ◽  
Author(s):  
Thomas R. Neu ◽  
Ute Kuhlicke ◽  
John R. Lawrence
Keyword(s):  
Nucleic Acid ◽  
Laser Scanning ◽  
Single Photon ◽  
Selective Excitation ◽  
Laser Scanning Microscopy ◽  
Scanning Microscopy ◽  
Excitation Wavelength ◽  
Laser Microscopy ◽  
Two Photon ◽  
Photon Excitation

ABSTRACT A major limitation for the use of two-proton laser scanning microscopy (2P-LSM) in biofilm and other studies is the lack of a thorough understanding of the excitation-emission responses of potential fluorochromes. In order to use 2P-LSM, the utility of various fluorochromes and probes specific for a range of biofilm constituents must be evaluated. The fluorochromes tested in this study included classical nucleic acid-specific stains, such as acridine orange (AO) and 4",6"-diamidino-2-phenylindole (DAPI), as well as recently developed stains. In addition, stains specific for biofilm extracellular polymeric substances (EPS matrix components) were tested. Two-photon excitation with a Ti/Sapphire laser was carried out at wavelengths from 760 to 900 nm in 10-nm steps. It was found that autofluorescence of phototrophic organisms (cyanobacteria and green algae) resulted in strong signals for the entire excitation range. In addition, the coenzyme F420-related autofluorescence of methanogenic bacteria could be used to obtain images of dense aggregates (excitation wavelength, 780 nm). The intensities of the emission signals for the nucleic acid-specific fluorochromes varied. For example, the intensities were similar for excitation wavelengths ranging from 780 to 900 nm for AO but were higher for a narrower range, 780 to 810 nm, for DAPI. In selective excitation, fading, multiple staining, and combined single-photon-two-photon studies, the recently developed nucleic acid-specific fluorochromes proved to be more suitable regardless of whether they are intended for living or fixed samples. Probes specific for proteins and glycoconjugates allowed two-photon imaging of polymeric biofilm constituents. Selective excitation-emission was observed for Calcofluor White M2R (780 to 800 nm) and SyproOrange (880 to 900 nm). In addition, fluor-conjugated concanavalin A lectins were examined and provided acceptable two-photon emission signals at wavelengths ranging from 780 to 800 nm. Finally, CellTracker, a fluorochrome suitable for long-term labeling of microbial eucaryote cells, was found to give strong emission at wavelengths ranging from 770 to 810 nm. If fluorochromes have the same two-photon excitation cross section, they are suitable for multiple staining and multichannel recording. Generally, if an appropriate excitation wavelength and fluorochrome were used, it was possible to obtain more highly resolved images for thick biofilm samples with two-photon laser microscopy than with conventional single-photon laser microscopy. Due to its potential for higher resolution in light-scattering tissue-like material, such as biofilms, and extremely localized excitation, 2P-LSM is a valuable addition to conventional confocal laser scanning microscopy with single-photon excitation. However, further development of the method and basic research are necessary to take full advantage of nonlinear excitation in studies of interfacial microbial ecology.

Sign in / Sign up

Export Citation Format

Share Document