SEM-CLSM correlation microscopy and its application to electrospun gelatin fibers

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Abstract

The most comprehensive information about microstructure of the sample can be obtained by combining different types of high-resolution microscopy. This combination turns out to be especially informative when measurements are carried out not only on the same image, but on the same area of the sample. This approach is called correlation microscopy. Typically, such experiments require careful preparation of the sample and transferring it between the two microscopes. The current work uses correlation microscopy which combines scanning electron microscopy (SEM) and confocal laser scanning microscopy (CLSM). Electrospun gelatin fibers deposited onto metallized glass are studied using these two methods. The possibility of using correlation analysis to combine images obtained by SEM and CLSM is demonstrated.

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About the authors

D. V. Bagrov

Lomonosov Moscow State University, Biological Department

Author for correspondence.
Email: bagrov@mail.bio.msu.ru
ORCID iD: 0000-0002-6355-7282

Cand. of Sci. (Physics and Mathematics), Leading Researcher

Russian Federation, Moscow

E. R. Pavlova

LOPUKHIN FRCC PCM

Email: bagrov@mail.bio.msu.ru
ORCID iD: 0000-0002-2511-7622

Cand. of Sci. (Physics and Mathematics), Researcher

Russian Federation, Moscow

A. S. Bogdanova

LOPUKHIN FRCC PCM; Moscow Institute of Physics and Technology

Email: bagrov@mail.bio.msu.ru
ORCID iD: 0000-0001-5369-8519

Junior Researcher

Russian Federation, Moscow; Dolgoprudny

A. M. Moysenovich

Lomonosov Moscow State University, Biological Department

Email: bagrov@mail.bio.msu.ru
ORCID iD: 0000-0001-5379-5829

Cand. of Sci. (Biological), Leading Researcher

Russian Federation, Moscow

T. V. Mitko

LOPUKHIN FRCC PCM; Moscow Institute of Physics and Technology

Email: bagrov@mail.bio.msu.ru
ORCID iD: 0000-0002-0107-1906

Cand. of Sci. (Biological), Junior Researcher

Russian Federation, Moscow; Dolgoprudny

A. A. Ramonova

Lomonosov Moscow State University, Biological Department

Email: bagrov@mail.bio.msu.ru
ORCID iD: 0000-0002-3081-4721

Junior Researcher

Russian Federation, Moscow

D. V. Klinov

LOPUKHIN FRCC PCM; Moscow Institute of Physics and Technology

Email: bagrov@mail.bio.msu.ru
ORCID iD: 0000-0001-8288-2198

Cand. of Sci. (Physics and Mathematics), Head of Laboratory

Russian Federation, Moscow; Dolgoprudny

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Supplementary files

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2. Fig.1. Design of the experiment. Electrospun fibres are arranged on a metal coating on glass, and a mark in the form of a cross is made on the coating

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3. Fig.2. Results of the correlation microscopy experiment. a, b – SEM images, yellow frames show the region of interest location; c – CLSM image; d – inverted SEM image prepared for correlation analysis; e – deformed image (d); f – deformed image (c). In panels e and f, a green triangle marks two fibres whose locations are markedly different, and red arrows mark a ribbon that is clearly visible by SEM and invis- ible to CLSM

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4. Fig.3. A pair of images obtained by CLSM (A, maximum intensity projection, image inverted) and SEM (B) prepared for correlation analysis. Reference points are shown in orange, red arrows indi- cate ribbons that are visible in the SEM study but not visible in the CLSM study. Image size 130 × 130 µm

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5. Fig.4. Comparison of fibre width data obtained from SEM and CLSM images

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Copyright (c) 2024 Bagrov D.V., Pavlova E.R., Bogdanova A.S., Moysenovich A.M., Mitko T.V., Ramonova A.A., Klinov D.V.

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