Comparative Evaluation of Common Fixatives on Histomorphological Preservation in Mouse Tissues Using Hematoxylin and Eosin Staining
DOI:
https://doi.org/10.54361/LJMR.20.2.47Keywords:
Histological preservation, Formalin fixation, Bouin’s fixation, Carnoy’s fixation, H&E stainingAbstract
Background: This study aimed to evaluate and compare the histological effects of three commonly used fixatives (10% neutral buffered formalin, Bouin’s solution, and Carnoy’s solution) on mouse liver, heart, and testis tissues.Methods: A total of twelve healthy adult male mice were used, with four mice allocated to each fixative group. From each animal, the liver, heart, and testis were surgically excised and fixed in the designated solutions for standard durations. Tissues were processed using an automated tissue processor, embedded in paraffin, sectioned at 3 μm thickness, and stained with hematoxylin and eosin (H&E). Histological sections were then evaluated microscopically for nuclear clarity, cytoplasmic preservation, staining quality, and structural integrity.Results: The results demonstrated notable differences among the fixatives. Bouin’s solution provided superior nuclear detail in testicular tissue but was associated with mild tissue hardening. Formalin exhibited optimal preservation of overall morphology, particularly in liver and heart sections, with minimal artifacts. Carnoy’s fixative offered sharp nuclear contrast but was associated with noticeable tissue shrinkage and distortion.Conclusion: These findings highlight that the choice of fixative significantly affects histological outcomes, and selection should be guided by tissue type and diagnostic objectives. The study underscores the importance of appropriate fixation in enhancing tissue preservation and microscopic interpretation.
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References
1. Cibas ES, Ducatman BS. Cytology: Diagnostic Principles and Clinical Correlates. 5th ed. Elsevier; 2020.
2. Nayar R, Wilbur DC. The Bethesda System for Reporting Cervical Cytology: Definitions, Criteria, and Explanatory Notes. 3rd ed. Springer; 2023.
3. Bishop JA. Molecular advances in cytopathology. J Clin Pathol. 2024;77(2):85-92. doi:10.1136/jclinpath2024-208544.
4. Alkali A, Hassan M, Yusuf T. Artifacts and fixation failures in histopathology: implications for diagnosis. Int J Pathol. 2024;16(1):12-20. doi:10.1016/j.ijpath.2024.01.003.
5. World Health Organization. Guidelines for Screening and Early Cancer Detection. Geneva: WHO; 2023.
6. Ahmed HG, Mohammed AI, Hussein MOM. A comparison study of histochemical staining after Carnoy's and formalin fixation. Sudan J Med Sci. 2011;5(4):285-290.
7. Ajileye O, Esan A. Standardization and usage of formalin in routine histopathology. J Histol Methods. 2022;15(3):120-130.
8. Ahmed S, Khan M, Al Hassan R. Limitations of Bouin’s solution for long-term tissue preservation. Int J Pathol. 2011;9(2):85-90.
9. Ananthanarayanan V, Pins MR, Meyer RE, Gann PH. Immunohistochemical assays in prostatic biopsies processed in Bouin’s fixative. J Clin Pathol. 2005;58(3):322-324.
10. Aziz R, Khan T, Malik S. Evaluation of spermatogenesis using Bouin’s fixative. Andrology Reports. 2023;10(1):45-53.
11. Bancroft JD, Gamble M. Theory and Practice of Histological Techniques. 6th ed. Elsevier; 2008.
12. Carson FL. Histotechnology: A Self-Instructional Text. 3rd ed. ASCP Press; 2009.
13. Dortbudak M, Eren F, Yilmaz A. Comparative analysis of fixatives on tissue morphology and staining. J Biomed Sci. 2024;32(2):105-113.
14. Fischer AH, Jacobson KA, Rose J, Zeller R. Hematoxylin and eosin staining of tissue and cell sections. Cold Spring Harb Protoc.
2008;2008(5):pdb.prot4986.
doi:10.1101/pdb.prot4986.
15. Howat WJ, Wilson BA. Tissue fixation and antigen preservation. Histopathology. 2014;65(2):200-210.
16. Pereira C, Sousa A, Martins M. Carnoy’s solution for nucleic acid preservation. Mol Pathol J.
2015;19(4):345-353.
17. Rowley JE, Chen L, Patel S. Tissue shrinkage effects in histology. J Clin Histol. 2020;27(3):78-86.
18. Singhal P, Gupta R, Verma N. Nuclear detail preservation in various fixatives. Diagn Pathol. 2016;11:42. doi:10.1186/s13000-016-0472-3.
19. Tian H, Li X, Wang Y. Preservation of chromosomes using Carnoy’s fixative. Cytogenet Cell Genet. 2024;18(1):55-63.
20. Thavarajar R, Mudinimannar VK, Elizabeth J, Rao UK, Ranganathan K. Chemical and physical basics of routine formalin fixation. J Oral Maxillofac Pathol.
2012;16(3):400-405. doi:10.4103/0973-
029X.102496.
21. Elzabal MHE, Ahmed HG, Abdelaziz MS, et al. Effect of fixative temperatures on histochemical staining. Sudan J Med Sci. 2013;8(3):147-150.
22. Dörtbudak MB, Demircioğlu M, Seker U,
Demircioğlu T. Comparison of different fixatives effects in histochemical stainings of peripheral nerve tissue. Cell Mol Biol (Noisy-le-grand).
2025;70(12):88-94. doi:10.14715/cmb/2024.70.12.12.
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Copyright (c) 2026 Rema A. Ashmila, Enass K. Ghwarsha, Najway Khaled Al-Ojli, Aziza S. Al-Suwaii, Fadwa Said Saleh Altarbaghia, Heba I. Al-Shalmani, Nouran J. Alshiky, Yousef M. Ali Hasen, Ahmed S Mikael, Salem Aldrsy, Mahmoud A. Alorib (Author)
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