Abstract
The use of semithin sections is important in histological and clinical diagnostic studies. However, since the plastic embedding media used for this purpose are hydrophobic, they limit the penetration of dyes into the tissues. In this study, it was aimed to compare the known epon solvents and acetone to provide more detail in the staining of semithin epon sections and to evaluate the toluidine blue-eosin double staining to be tried for the first time. For this purpose, 1μm thick semithin sections were taken from epon blocks of rat brain, small intestine and pancreatic tissue samples with an ultramicrotome. Before staining, three different etchings were applied to the epon sections with sodium hydroxide, periodic acid and acetone in order to facilitate the penetration of the dyes. Epon was not removed in one set of sections. Sections with staining of toluidine blue and toluidine blue-eosin were evaluated under the light microscope. The toluidine blue staining results of three tissues after etching with acetone were evaluated as much more successful than the stainings after alkali and acid etching and staining without etching. It was observed that it provided clearer images, especially in the pancreas. Positive results were obtained in brain and pancreatic tissues with toluidine blue-eosin staining. Since successful results were obtained, it was concluded that acetone can be used routinely as an alternative to other solvents in the removal of epon, and toluidine blue-eosin double staining can be used in the evaluation of epon sections.
References
- 1. Hayat M.A. Staining of semithin sections. In: Hayat MA (ed). Principles and Techniques of Electron Microscopy. 4th edition. Cambridge: Cambridge University Press; 2000. 360-6.
- 2. Robinson G, Gray T. Electron microscopy: Practical procedures. In: Bancroft JD, Stevens A (eds). Theory and Practice of Histological Techniques. 4th edition. Edinburgh: Churchill Livingstone; 1996. 585-626.
- 3. Woods AE, Stirling JW. Transmission electron microscopy. In: Suvarna K, Layton C, Bancroft JD (eds). Bancroft’s Theory and Practice of Histological Techniques. 7th edition. China: Churchill Livingstone; 2013. 493-538.
- 4. Horobin RW. Staining plastic sections: a rewiew of problems, explanations and possible solutions. J Microsc 1983;131(2):173-86.
- 5. Trump BF, Smuckler EA, Benditt EP. A method for staining epoxy sections for light microscopy. J Ultrastruct Res 1961;5:343-48.
- 6. Iwadare T, Arai T. Staining etched epoxy resin sections for light microscopy. Biotech Histochem 1995;70(2):53-6.
- 7.Van Reempts J, Borgers M. A simple polychrome stain forconventionally fixed epon-embedded tissues. Stain Technol 1975;50(1):19-23.
- 8.Tolivia J, Navarro A, Tolivia D. Polychromatic staining ofepoxy semithin sections: a new and simple method. Histochemistry 1994;101(1):51-5.
- 9.Cai Z, Manavis J, Cash K, Thompson PD, Blumbergs PC.Immunohistochemical staining of epoxy resin sections of peripheral nerve. Appl Immunohistochem Mol Morphol 2005;13(3): 292-4.
- 10.Zhai XY, Kristoffersen IB, Christensen EI. Immunocytochemistry of renal membrane proteins on epoxysections. Kidney Int 2007;72(6):731-5.
- 11.Krueger SK, Phillips DE, Frederick MM, Johnson RK. Diaminobenzidine as a myelin stain in semithin plastic sections. Biotech Histochem 1999;74(2):105-9.
- 12.Haraguchi CM, Yokota S. Immunofluorescence technique for100-nm-thick semithin sections of epon-embedded tissues. Histochem Cell Biol 2002;117(1):81-5.
- 13.Iwadare T, Harada E, Yoshino S, Arai T. A solution forremoval of resin from epoxy sections. Stain Technol 1990;65(4):205-9.
- 14.Erenpreĭsa EA, Enkuzens AK. Improved method of staining semithin sections with toluidine blue. Arkh Patol 1980;42(8):82-3.
- 15.Roberts IM, Hutcheson AM. Handling and staining epoxy resinsections for light microscopy. J Microsc 1975;103(1):121-6.
- 16.Tzitsikas H, Rdzok EJ, Vatter AE. Staining residual lipids in ultrathin sections of tissues embedded in polyester resin. StainTechnol 1962;37(5):299-301.
Abstract
Yarı ince kesitlerin kullanımı histolojik ve klinik tanı çalışmalarında önemli bir yere sahiptir. Fakat bu amaçla kullanılan plastik gömme ortamları hidrofobik oldukları için boyaların dokulara girişini sınırlandırmaktadırlar. Bu çalışmada; yarı ince epon kesitlerin boyanmasında daha çok detay sağlamak üzere bilinen epon solventleri ile asetonun karşılaştırılması ve daha önce denenmemiş toluidine blue-eozin ikili boyamasının değerlendirilmesi amaçlanmıştır. Çalışmada sıçan beyin, ince bağırsak ve pankreas doku örneklerine ait epon bloklardan ultramikrotom ile 1μm kalınlığında yarı ince kesitler alındı. Boyama öncesi epon kesitlere boyaların penetrasyonunu kolaylaştırmak amacı ile sodyum hidroksit, periyodik asit ve aseton ile üç ayrı etching uygulandı. Bir grup kesitte epon uzaklaştırılmadı. Toluidine blue ve toluidine blue-eozin ikili boyaması uygulanan kesitler ışık mikroskobunda değerlendirildi. Aseton ile etching sonrası üç dokuya ait toluidine blue boyama sonuçları, alkali ve asit etchingi sonrası boyamalara ve etching uygulanmadan yapılan boyamalara göre çok daha başarılı olarak değerlendirildi. Özellikle pankreasta daha berrak görüntüler sayesinde daha iyi detay sağladığı görüldü. Toluidine blue-eozin boyaması ile beyin ve pankreas dokularında olumlu sonuçlar elde edildi. Eponun uzaklaştırılması amacıyla ilk kez denenen aseton ile başarılı sonuçlar elde edildiği için diğer solventlere alternatif olarak asetonun rutinde kullanılabileceği düşünüldü. Yine ilk kez denenen toluidine blue-eozin ikili boyamasından epon kesitlerin değerlendirilmesinde yararlanılabileceği sonucuna varıldı.
Keywords
References
- 1. Hayat M.A. Staining of semithin sections. In: Hayat MA (ed). Principles and Techniques of Electron Microscopy. 4th edition. Cambridge: Cambridge University Press; 2000. 360-6.
- 2. Robinson G, Gray T. Electron microscopy: Practical procedures. In: Bancroft JD, Stevens A (eds). Theory and Practice of Histological Techniques. 4th edition. Edinburgh: Churchill Livingstone; 1996. 585-626.
- 3. Woods AE, Stirling JW. Transmission electron microscopy. In: Suvarna K, Layton C, Bancroft JD (eds). Bancroft’s Theory and Practice of Histological Techniques. 7th edition. China: Churchill Livingstone; 2013. 493-538.
- 4. Horobin RW. Staining plastic sections: a rewiew of problems, explanations and possible solutions. J Microsc 1983;131(2):173-86.
- 5. Trump BF, Smuckler EA, Benditt EP. A method for staining epoxy sections for light microscopy. J Ultrastruct Res 1961;5:343-48.
- 6. Iwadare T, Arai T. Staining etched epoxy resin sections for light microscopy. Biotech Histochem 1995;70(2):53-6.
- 7.Van Reempts J, Borgers M. A simple polychrome stain forconventionally fixed epon-embedded tissues. Stain Technol 1975;50(1):19-23.
- 8.Tolivia J, Navarro A, Tolivia D. Polychromatic staining ofepoxy semithin sections: a new and simple method. Histochemistry 1994;101(1):51-5.
- 9.Cai Z, Manavis J, Cash K, Thompson PD, Blumbergs PC.Immunohistochemical staining of epoxy resin sections of peripheral nerve. Appl Immunohistochem Mol Morphol 2005;13(3): 292-4.
- 10.Zhai XY, Kristoffersen IB, Christensen EI. Immunocytochemistry of renal membrane proteins on epoxysections. Kidney Int 2007;72(6):731-5.
- 11.Krueger SK, Phillips DE, Frederick MM, Johnson RK. Diaminobenzidine as a myelin stain in semithin plastic sections. Biotech Histochem 1999;74(2):105-9.
- 12.Haraguchi CM, Yokota S. Immunofluorescence technique for100-nm-thick semithin sections of epon-embedded tissues. Histochem Cell Biol 2002;117(1):81-5.
- 13.Iwadare T, Harada E, Yoshino S, Arai T. A solution forremoval of resin from epoxy sections. Stain Technol 1990;65(4):205-9.
- 14.Erenpreĭsa EA, Enkuzens AK. Improved method of staining semithin sections with toluidine blue. Arkh Patol 1980;42(8):82-3.
- 15.Roberts IM, Hutcheson AM. Handling and staining epoxy resinsections for light microscopy. J Microsc 1975;103(1):121-6.
- 16.Tzitsikas H, Rdzok EJ, Vatter AE. Staining residual lipids in ultrathin sections of tissues embedded in polyester resin. StainTechnol 1962;37(5):299-301.
Details
| Primary Language | Turkish |
|---|---|
| Subjects | Anatomy |
| Journal Section | Research Article |
| Authors | |
| Publication Date | June 9, 2023 |
| Acceptance Date | May 29, 2023 |
| Published in Issue | Year 2023 Volume: 49 Issue: 1 |
Cite
Cited By
Journal of Uludag University Medical Faculty is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.