Attenuation of the Early Events of α-Synuclein Aggregation: A Fluorescence Correlation Spectroscopy and Laser Scanning Microscopy Study in the Presence of Surface-Coated Fe3O4 Nanoparticles

Langmuir ◽  
2015 ◽  
Vol 31 (4) ◽  
pp. 1469-1478 ◽  
Author(s):  
Nidhi Joshi ◽  
Sujit Basak ◽  
Sangeeta Kundu ◽  
Goutam De ◽  
Anindita Mukhopadhyay ◽  
...  
Keyword(s):  
Fe3o4 Nanoparticles ◽  
Fluorescence Correlation Spectroscopy ◽  
Laser Scanning ◽  
Microscopy Study ◽  
Correlation Spectroscopy ◽  
Laser Scanning Microscopy ◽  
Scanning Microscopy ◽  
Fluorescence Correlation

scholarly journals Fluorescence Correlation Spectroscopy Combined with Multiphoton Laser Scanning Microscopy—A Practical Guideline

2021 ◽  
Vol 11 (5) ◽  
pp. 2122
Author(s):  
Jeemol James ◽  
Jonas Enger ◽  
Marica B. Ericson
Keyword(s):  
Biomedical Research ◽  
Fluorescence Correlation Spectroscopy ◽  
Laser Power ◽  
Laser Scanning ◽  
Correlation Spectroscopy ◽  
Operating Conditions ◽  
Laser Scanning Microscopy ◽  
Scanning Microscopy ◽  
Fluorescence Correlation ◽  
Practical Guideline

Multiphoton laser scanning microscopy (MPM) has opened up an optical window into biological tissues; however, imaging is primarily qualitative. Cell morphology and tissue architectures can be clearly visualized but quantitative analysis of actual concentration and fluorophore distribution is indecisive. Fluorescence correlation spectroscopy (FCS) is a highly sensitive photophysical methodology employed to study molecular parameters such as diffusion characteristics on the single molecule level. In combination with laser scanning microscopy, and MPM in particular, FCS has been referred to as a standard and highly useful tool in biomedical research to study diffusion and molecular interaction with subcellular precision. Despite several proof-of-concept reports on the topic, the implementation of MPM-FCS is far from straightforward. This practical guideline aims to clarify the conceptual principles and define experimental operating conditions when implementing MPM-FCS. Validation experiments in Rhodamine solutions were performed on an experimental MPM-FCS platform investigating the effects of objective lens, fluorophore concentration and laser power. An approach based on analysis of time-correlated single photon counting data is presented. It is shown that the requirement of high numerical aperture (NA) objective lenses is a primary limitation that restricts field of view, working distance and concentration range. Within these restrictions the data follows the predicted theory of Poisson distribution. The observed dependence on laser power is understood in the context of perturbation on the effective focal volume. In addition, a novel interpretation of the effect on measured diffusion time is presented. Overall, the challenges and limitations observed in this study reduce the versatility of MPM-FCS targeting biomedical research in complex and deep tissue—being the general strength of MPM in general. However, based on the systematic investigations and fundamental insights this report can serve as a practical guide and inspire future research, potentially overcoming the technical limitations and ultimately allowing MPM-FCS to become a highly useful tool in biomedical research.

scholarly journals Bacterial Colonization and Proliferation in Furcal Perforations Repaired by Different Materials: A Confocal Laser Scanning Microscopy Study

2021 ◽  
Vol 11 (8) ◽  
pp. 3403
Author(s):  
Shlomo Elbahary ◽  
Sohad Haj Yahya ◽  
Cemre Koç ◽  
Hagay Shemesh ◽  
Eyal Rosen ◽  
...  
Keyword(s):  
Confocal Laser Scanning Microscopy ◽  
Laser Scanning ◽  
Bacterial Colonization ◽  
Microscopy Study ◽  
Laser Scanning Microscopy ◽  
Confocal Laser Scanning ◽  
Scanning Microscopy ◽  
Confocal Laser ◽  
Repair Material ◽  
Dentinal Tubules

Following furcal perforation, bacteria may colonize the defect and cause inflammation and periodontal destruction. This study used confocal laser scanning microscopy (CLSM) to evaluate Enterococcus faecalis colonization and proliferation in furcal perforations repaired with different materials. Furcal perforations created in 55 extracted human mandibular molars were repaired using either MTA-Angelus, Endocem, or Biodentine and coronally subjected to E. faecalis suspension for 21 days. The specimens were then stained using a LIVE/DEAD Viability Kit and visualized by CLSM. The minimum and maximum depths of bacterial penetration into the dentinal tubules were 159 and 1790 μM, respectively, with a mean of 713 μM. There were significantly more live than dead bacteria inside the dentinal tubules (p = 0.0023) in all groups, and all three repair materials exhibited a similarly sized stained area (p = 0.083). However, there were significant differences in the numbers of dead bacteria at the circumference of the perforation defect (p = 0.0041), with a significantly higher ratio of live to dead bacteria in the MTA-Angelus group (p = 0.001). Following perforation repair, bacteria may colonize the interface between the repair material and dentin and may penetrate through the dentinal tubules. The type of repair material has a significant effect on the viability of the colonizing bacteria.

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