Comparing two methods of plastination and glycerin preservation to study skeletal system after Alizarin red-Alcian blue double staining

Authors
Mohsen M Setayesh
Ebrahim Esfandiari
Abbas A Rabiei
Mahsa S Hanaei
Bahman Rashidi
Department of Anatomical Sciences, School of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran
Abstract
Background: Plastination is a new method of preserving tissue samples for a long time. This study aimed to compare the new plastination technique with the conventional preservative method in glycerin for fetus skeleton tissues and young rats dyed by Alizarin red- Alcian blue double staining.
Materials and Methods: In this study, 4 groups of 1-day, 3-day, 12-day and mature rats were selected and, after being anesthetized and slaughtered, their skin was completely removed. In Alizarin red- Alcian blue double staining method, first the samples were fixed in 95% ethanol and then their cartilages were dyed by 0.225% Alcian blue solution; after that, they were cleared in 1% KOH. Then, the bones were dyed in 0.003% Alizarin red solution and finally the tissue was decolorized in 95% ethanol. In each group, half of the samples were preserved by the conventional method in a glycerin container and the other half were plastinated.
Results: In the present study, the samples preserved by plastination technique were dry, odorless, indecomposable and tangible. Quality of coloring had an inverse relationship with rats' age. Transparency of the plastinated samples had also an inverse relationship with rats' age. Therefore, skeletal tissue of younger rats had higher quality and transparency in both preservation methods (glycerin and plastination).
Conclusion: This study showed that plastination technique was an appropriate method in comparison with glycerin preservation, which conserved skeletal tissue of fetus and young rats colored by Alizarin red- Alcian blue double staining. And the final result was that plastination technique can generate dry, odorless, indecomposable and tangible samples.

Keywords

1. Leena Pramod K, Vaswani VR, Bind hu S. Museum preservation of skeleton of fetus and small vertebrates. Rec Res Sci Tech 2011;3:54-8.  Back to cited text no. 1
    
2. Culling CFA. Museum technique. Handbook of histopathological techniques (including museum technique), 2nd Ed., London: Butterworths; 1963. p. 523-38.  Back to cited text no. 2
    
3. Cortes Delgado N, Perez Torres J, Mario Hoyos J. Staining procedure of cartilage and skeleton in adult bats and rodents. Int J Morphol 2009;27:1163-7.  Back to cited text no. 3
    
4. Alden BD. A note on the staining of the skeleton of cleared specimens with Alizarin red. Stain Technol 1962;1:123-4.  Back to cited text no. 4
    
5. Inouye M. Differential staining of cartilage and bone in fetal mouse skeleton by Alcian blue and Alizarin red. Congenit Anom (Kyoto) 1976;16:171-3.  Back to cited text no. 5
    
6. Erdodon D, Kadiodlu D, Peker T. Visualization of the fetal skeletal system by double staining with alizarin red and alcian blue. Gazi Medical Journal 1995;6:55-8.  Back to cited text no. 6
    
7. Thompsett DH. Cleared anatomical specimens. Anatomical Techniques. 2 nd ed. Chapter 35. Edinburgh: E. & S. Livingstone. 1970. p. 248-58.  Back to cited text no. 7
    
8. von Hagens G, Tiederman K, Kriz W. The current potential of plastination. Anat Embryol (Berl) 1987;175:411-21.  Back to cited text no. 8
    
9. Gao H, Liu J, Yu S, Sui H. A new polyester technique for sheet plastination. J Int Soc Plastination 2006;21:7-10.  Back to cited text no. 9
    
10. De Roberti EM. Alcian blue - alizarin red skeletal staining. 2003. Available from: http://www.hhmi.ucla.edu/derobertis/-Protocols- Mouse Protocols-. [Last updated on 2011 Jan 22.  Back to cited text no. 10
    
11. Dingerkus G, Uhler LD. Enzyme clearing of alcian blue stained whole small vertebrates' dor demonstration of cartilage. Stain Technol 1997;52:229-32.  Back to cited text no. 11
    
12. Potthof T. Clearing and staining techniques. In: Moser HD, editor. Ontogeny and systematic of fishes. vol. 1. KS, USA: Special publication-American society of Ichthyologists and Herpetologist, Lawrence; 1984. p. 35-7.  Back to cited text no. 12
    
13. Wang Y, Xiao R, Yong F, Karim Bo, Lacovelli AJ, Cai J. Abnormalities in cartilage and bone development in the Apert syndrome FGFR2+S252W mouse. Development 2005;132:3537-48.  Back to cited text no. 13
    
14. Ranjbar R, Vatanchian M, Mohammadian B, Mayahi M. Chronological study of chicken embryo wing and leg skeleton development by alizarin red- alcian blue double staining technique. J Iranian Biology 1385;19:164-79.  Back to cited text no. 14
    
15. Rabiei AA. Fabrication of polymer production of the whole human body with mass plastination method [PHD Thesis]. Isfahan: Isfahan University of Medical Science; 2003.  Back to cited text no. 15
    
16. Rabiei AA., Asadi MH, Esfandiari E, Taghipour M, Bahadoran H, Setayesh M. Prepration of flexible plastinated sheets of human brain by P87 polyester. J Isfahan Med Sch 2011;28:1961-6.  Back to cited text no. 16
    
17. Von Hagen G. Preservation by plastination. J Int Soc Plastination 1994;7:5-19.  Back to cited text no. 17
    
18. Henry RW, Lotorre R. Polyester plastination of biological tissue. J Int Soc Plastination 2007;22:59-68.  Back to cited text no. 18
    
19. Haffajee MR. Plastination of a cleared fetus to show vascularization of ossifying bone. J Int Soc Plastination 1996;10:26-7.  Back to cited text no. 19
    
20. Steinke H, Rabi S, Saito T. Staining body slices before and after plastination. Eur J Anat 2008;12:51-5.  Back to cited text no. 20
Advanced Biomedical Research
Volume 2013, March
March 2013
Pages 1-4