Glutaraldehyde-enhanced autofluorescence as a general tool for 3D morphological imaging

Miika Niemeläinen; Anna-Mari Haapanen-Saaristo; Leena M Koskinen; Josef Gullmets; Emilia Peuhu; Annika Meinander; Sara Calhim; Ilkka Paatero

doi:10.1242/bio.060428

Glutaraldehyde-enhanced autofluorescence as a general tool for 3D morphological imaging

Miika Niemeläinen
, Anna-Mari Haapanen-Saaristo
, Leena M Koskinen
, Josef Gullmets
, Emilia Peuhu
, Annika Meinander
, Sara Calhim
, Ilkka Paatero

Research output: Contribution to journal › Article › Scientific › peer-review

1 Citation (Scopus)

126 Downloads (Pure)

Abstract

Routine histochemical techniques are capable of producing vast amount of information from diverse sample types, but these techniques are limited in their ability to generate 3D information. Autofluorescence imaging can be used to analyse samples in 3D but it suffers from weak/low signal intensities. Here, we describe a simple chemical treatment with glutaraldehyde to enhance autofluorescence for 3D fluorescence imaging and to generate detailed morphological images on whole-mount samples. This methodology is straightforward and cost-effective to implement, suitable for a wide range of organisms and sample types. Furthermore, it can be readily integrated with standard confocal and fluorescence microscopes for analysis. This approach has the potential to facilitate the analysis of biological 3D structures and research in developmental biology, including studies on model and non-model organisms.

Original language	English
Article number	bio060428
Journal	Biology Open
Volume	13
Issue number	11
DOIs	https://doi.org/10.1242/bio.060428
Publication status	Published - Nov 2024
MoE publication type	A1 Journal article-refereed

Funding

We acknowledge Zebrafish Core and Cell Imaging Core (Turku Bioscience Centre, University of Turku and \u00C5bo Akademi University), the University of Helsinki Drosophila core facility (Hi-Fly) and the \u00C5bo Akademi University Fly Unit, all supported by Biocenter Finland, for expertise, assistance and providing research infrastructure. This research was supported by InFLAMES Flagship Programme of the Academy of Finland (decision numbers: 337531, 357911). Business Finland (1718/31/2021, I.P.), FinnFOTO (20220515, I.P.), the \u00C5bo Akademi University Foundation (AM), Sigrid Jus\u00E9lius Foundation (E.P), Academy of Finland research fellowship (323096, E.P.), the Hospital District of Southwest Finland (11083, E.P.), Turku Doctoral Program of Molecular Medicine (L.M.K.) Open Access funding provided by University of Turku: Turun Yliopisto. Deposited in PMC for immediate release. Business Finland (1718/31/2021, I.P.), FinnFOTO (20220515, I.P.), the \u00C5bo Akademi University Foundation (AM), Sigrid Juse\u0301lius Foundation (E.P), Academy of Finland research fellowship (323096, E.P.), the Hospital District of Southwest Finland (11083, E.P.), Turku Doctoral Program of Molecular Medicine (L.M.K.) Open Access funding provided by University of Turku: Turun Yliopisto. Deposited in PMC for immediate release. We acknowledge Zebrafish Core and Cell Imaging Core (Turku Bioscience Centre, University of Turku and \u00C5bo Akademi University), the University of Helsinki Drosophila core facility (Hi-Fly) and the \u00C5bo Akademi University Fly Unit, all supported by Biocenter Finland, for expertise, assistance and providing research infrastructure. This research was supported by InFLAMES Flagship Programme of the Academy of Finland (decision numbers: 337531, 357911). Strains were provided by the CGC, which is funded by NIH Office of Research Infrastructure Programs (P40 OD010440). Mixed stage cultures of C. elegans [N2 (wild-type), CB4856 (C. elegans wild isolate), CB769 (bli-1(e769) II) and CB128 (dpy-10(e128) II] were maintained using standard protocols (Stiernagle, 2006). For fixation and staining, the worms were washed off from NGM plates. The worms were fixed with 2.5% glutaraldehyde in PBS for overnight at RT. After several washes with PBSTw, the worms were mounted into low-melting point agarose (0.7%) on glass-bottom dishes for imaging with 3i CSU-W1 spinning disk, 40\u00D7 Zeiss LD C-Apochromat WI or 40\u00D7 Zeiss LD Plan-Neofluar objective and excitation at 561 nm.

Keywords

Imaging, Three-Dimensional/methods
Glutaral/chemistry
Animals
Optical Imaging/methods
Microscopy, Fluorescence/methods
Microscopy, Confocal/methods
Humans

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10.1242/bio.060428Licence: CC BY

bio060428Final published version, 4.98 MBLicence: CC BY

https://urn.fi/URN:NBN:fi-fe20241216102790

Cite this

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abstract = "Routine histochemical techniques are capable of producing vast amount of information from diverse sample types, but these techniques are limited in their ability to generate 3D information. Autofluorescence imaging can be used to analyse samples in 3D but it suffers from weak/low signal intensities. Here, we describe a simple chemical treatment with glutaraldehyde to enhance autofluorescence for 3D fluorescence imaging and to generate detailed morphological images on whole-mount samples. This methodology is straightforward and cost-effective to implement, suitable for a wide range of organisms and sample types. Furthermore, it can be readily integrated with standard confocal and fluorescence microscopes for analysis. This approach has the potential to facilitate the analysis of biological 3D structures and research in developmental biology, including studies on model and non-model organisms.",

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AU - Koskinen, Leena M

AU - Gullmets, Josef

AU - Peuhu, Emilia

AU - Meinander, Annika

AU - Calhim, Sara

AU - Paatero, Ilkka

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KW - Animals

KW - Optical Imaging/methods

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Glutaraldehyde-enhanced autofluorescence as a general tool for 3D morphological imaging

Abstract

Funding

Keywords

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