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. 2022 Dec;36(12):e24794.
doi: 10.1002/jcla.24794. Epub 2022 Nov 28.

Effects of six kinds of sperm staining methods on human sperm size and evaluation of their staining effects

Yuan-Hua Xu  1 , Jin-Chun Lu  1 , Shan-Shan Tang  1
Affiliations

Effects of six kinds of sperm staining methods on human sperm size and evaluation of their staining effects

Yuan-Hua Xu et al. J Clin Lab Anal. 2022 Dec.

Abstract

Background: Large- and small-headed sperm are common morphological abnormalities. If different sperm staining methods affect sperm size, they will make a difference in the accuracy of sperm morphological analysis results. In this case, the normal reference values of sperm head parameters for different staining methods should be established.

Methods: Six sperm staining methods, including Papanicolaou, Diff-Quik, Shorr, Hematoxylin-eosin (HE), Wright, and Wright-Giemsa staining, were used to stain the sperm smears of 25 semen samples, respectively. Sperm head parameter's length (L), width (W), area (A), perimeter, acrosomal area (Ac), and the derived values L/W and Ac/A of 2500 sperm (100 for each specimen) per staining method were measured by a computer-aided sperm morphological analysis system.

Results: The highest sperm head length and width were observed with the Wright-Giemsa and Wright staining, followed by the Diff-Quik. The lowest sperm head length and width were observed with the Papanicolaou staining, and the sperm head length and width of HE and Shorr staining were between those of Papanicolaou and Diff-Quik staining. There was the same trend in changes in sperm head area and perimeter. Diff-Quik and Shorr staining could clearly distinguish acrosome and nucleus, followed by HE staining, whereas the boundary between acrosome and nucleus was not evident in Papanicolaou, Wright, and Wright-Giemsa staining.

Conclusion: Different staining methods influence sperm size, and the normal reference values of sperm head parameters of each staining method should be established. Diff-Quik and Shorr staining may be suitable methods for routine sperm morphological analysis.

Keywords: human sperm; morphological analysis; sperm size; staining method; standardization.

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Conflict of interest statement

All authors declare no competing interests.

Figures

FIGURE 1
FIGURE 1
Sperm images for Diff‐Quik staining.
FIGURE 2
FIGURE 2
Sperm images for Shorr staining.
FIGURE 3
FIGURE 3
Sperm images for Papanicolaou staining.
FIGURE 4
FIGURE 4
Sperm images for Hematoxylin–eosin staining.
FIGURE 5
FIGURE 5
Sperm images for Wright staining.
FIGURE 6
FIGURE 6
Sperm images for Wright‐Giemsa staining.
See this image and copyright information in PMC

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References

    1. Gacem S, Catalán J, Yánez‐Ortiz I, Soler C, Miró J. New sperm morphology analysis in equids: Trumorph® vs eosin‐nigrosin stain. Vet Sci. 2021;8:79. - PMC - PubMed
    1. Lu JC. Analysis of sperm morphology: yes or no? Nat J Androl. 2013;19:291‐295. - PubMed
    1. Ammar O, Mehdi M, Muratori M. Teratozoospermia: its association with sperm DNA defects, apoptotic alterations, and oxidative stress. Andrology. 2020;8:1095‐1106. - PubMed
    1. Singh S, Sharma S, Jain M, Chauhan R. Importance of papanicolaou staining for sperm morphologic analysis: comparison with an automated sperm quality analyzer. Am J Clin Pathol. 2011;136:247‐251. - PubMed
    1. Fucic A, Maric T, Vicic Bockor V, Jezek D. In vivo acridine orange human zoospermia staining‐a new perspective for RNA detection and zoospermia morphology evaluation. Anat Histol Embryol. 2021;50:102‐107. - PubMed