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Impact of Fixation Protocols on Elemental Content in Human Sperm

Tufoni, Cristina ; Gianoncelli, Alessandra ; Žižić, Milan ; Sala, Simone LU ; Kahnt, Maik LU orcid ; De Rocco, Daniela ; Luppi, Stefania ; Ricci, Giuseppe and Pascolo, Lorella (2025) In X-Ray Spectrometry
Abstract

X-ray Fluorescence microscopy is a sophisticated advanced technique which can provide information regarding the concentration and the distribution of chemical elements within biological samples. It is increasingly used in the study of cells and tissues as it does not require the use of staining or fluorescent probes. For XRF measurements, sample preparation is a key step in the analytical process that affects all aspects of the analysis. Cells should be maintained in conditions that closely resemble physiological environments, but this can be challenging, especially under vacuum conditions. While numerous fixation protocols exist, finding one that works best for all cell types remains difficult. This study focuses on human spermatozoa,... (More)

X-ray Fluorescence microscopy is a sophisticated advanced technique which can provide information regarding the concentration and the distribution of chemical elements within biological samples. It is increasingly used in the study of cells and tissues as it does not require the use of staining or fluorescent probes. For XRF measurements, sample preparation is a key step in the analytical process that affects all aspects of the analysis. Cells should be maintained in conditions that closely resemble physiological environments, but this can be challenging, especially under vacuum conditions. While numerous fixation protocols exist, finding one that works best for all cell types remains difficult. This study focuses on human spermatozoa, as they differ from other cell types in size, structure, and function. Furthermore, in human sperm, the presence and the balance of various chemical elements are crucial to their efficiency and success in reproduction. Our work aims at investigating the most suitable protocol for spermatozoa fresh cell fixation for XRF analyses at different energies; in particular, paraformaldehyde (PFA) 4% and methanol fixation are compared through XRF analyses at low and hard X-ray energy excitation, revealing light and trace elements. Although PFA is the most widely used chemical fixative for biological sample preparation, our results suggest that methanol is an excellent alternative for fixing human sperm for X-ray microscopy as it better preserves their elemental content while still maintaining an acceptably good morphology.

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author
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organization
publishing date
type
Contribution to journal
publication status
epub
subject
keywords
cell fixation, human sperm, methanol, PFA, X-ray Fluorescence
in
X-Ray Spectrometry
publisher
John Wiley & Sons Inc.
external identifiers
  • scopus:105013800004
ISSN
0049-8246
DOI
10.1002/xrs.70028
language
English
LU publication?
yes
additional info
Publisher Copyright: © 2025 John Wiley & Sons Ltd.
id
c50bbe6b-5b3c-461a-997a-d76e1d46e649
date added to LUP
2025-09-03 09:37:10
date last changed
2025-09-03 12:54:52
@article{c50bbe6b-5b3c-461a-997a-d76e1d46e649,
  abstract     = {{<p>X-ray Fluorescence microscopy is a sophisticated advanced technique which can provide information regarding the concentration and the distribution of chemical elements within biological samples. It is increasingly used in the study of cells and tissues as it does not require the use of staining or fluorescent probes. For XRF measurements, sample preparation is a key step in the analytical process that affects all aspects of the analysis. Cells should be maintained in conditions that closely resemble physiological environments, but this can be challenging, especially under vacuum conditions. While numerous fixation protocols exist, finding one that works best for all cell types remains difficult. This study focuses on human spermatozoa, as they differ from other cell types in size, structure, and function. Furthermore, in human sperm, the presence and the balance of various chemical elements are crucial to their efficiency and success in reproduction. Our work aims at investigating the most suitable protocol for spermatozoa fresh cell fixation for XRF analyses at different energies; in particular, paraformaldehyde (PFA) 4% and methanol fixation are compared through XRF analyses at low and hard X-ray energy excitation, revealing light and trace elements. Although PFA is the most widely used chemical fixative for biological sample preparation, our results suggest that methanol is an excellent alternative for fixing human sperm for X-ray microscopy as it better preserves their elemental content while still maintaining an acceptably good morphology.</p>}},
  author       = {{Tufoni, Cristina and Gianoncelli, Alessandra and Žižić, Milan and Sala, Simone and Kahnt, Maik and De Rocco, Daniela and Luppi, Stefania and Ricci, Giuseppe and Pascolo, Lorella}},
  issn         = {{0049-8246}},
  keywords     = {{cell fixation; human sperm; methanol; PFA; X-ray Fluorescence}},
  language     = {{eng}},
  month        = {{06}},
  publisher    = {{John Wiley & Sons Inc.}},
  series       = {{X-Ray Spectrometry}},
  title        = {{Impact of Fixation Protocols on Elemental Content in Human Sperm}},
  url          = {{http://dx.doi.org/10.1002/xrs.70028}},
  doi          = {{10.1002/xrs.70028}},
  year         = {{2025}},
}