A rapid pathological preparation method for composite material observation_Chinese Journal of Reparative and Reconstructive Surgery

Authors：

ZHANG Yi ¹ , LIANG Yan ¹ , DA Lincui ² ,  XIE Huiqi ²

1. Research Core Facility of West China Hospital, Sichuan University, Chengdu Sichuan, 610041, P.R.China;
2. Laboratory of Stem Cell and Tissue Engineering, West China Hospital, Sichuan University, Chengdu Sichuan, 610041, P.R.China;

Corresponding?author：

XIE Huiqi, Email: xiehuiqi@163.com

Keywords：

Composite material; frozen section; eosin staining; sealing

DOI：

10.7507/1002-1892.201806094

Video：

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Objective To explore a simple and rapid pathological slices method to observe the porous structure and the composition distribution of composite materials. Methods Taking polyurethane/small intestinal submucosa (PU/SIS) composite as an example, PU/SIS was OCT-embedded and sliced into sections by frozen section technology, after which general observation of the section integrity was carried out. After dyed with water-soluble eosin in alcoholic solution, the staining effect and the porous structure of the composite were observed under light field microscope. Sections were sealed with five different sealing methods. Group A: sealing piece using glycerogelatin method; group B: anhydrous alcohol dehydration→transparency using TO transparent reagent→sealing piece using neutral quick drying glue; group C: color separation using deionized water→air-drying→sealing piece using neutral quick drying glue; group D: air-drying→transparency using TO transparent reagent→sealing piece using neutral quick drying glue; group E: air-drying→sealing piece using neutral quick drying glue. Then, the morphology and the components distribution of the composite were observed under light field microscope, and the simple and feasible method was selected as optimum method. Results From general observation, the frozen section of the PU/SIS composite, which was 6 μm in thickness, was complete and continuous. Although the outline of the material and the porous structure in the sections could be observed clearly under light field microscope, the two components still could not be identified by using eosin staining method. After sealing piece, the material components in groups A, B, and C still could not be identified or be dissolved and deformed; the morphology of the material in groups D and E were preserved and the two components in the composite were clearly visible. Conclusion The morphology and the components distribution of PU/SIS frozen sections can be characterized after soluble eosin staining and neutral quick drying glue sealing.

Citation： ZHANG Yi, LIANG Yan, DA Lincui, XIE Huiqi. A rapid pathological preparation method for composite material observation. Chinese Journal of Reparative and Reconstructive Surgery, 2019, 33(1): 80-84. doi: 10.7507/1002-1892.201806094 Copy

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2.	Kuppan P, Sethuraman S, Krishnan UM. PCL and PCL-gelatin nanofibers as esophageal tissue scaffolds: optimization, characterization and cell-matrix interactions. J Biomed Nanotechnology, 2013, 9(9): 1540-1555.
3.	Jwo SC, Chin CH, Tang SJ, et al. Tubular scaffolds of gelatin and poly (ε-caprolactone)-block-poly (γ-glutamic acid) blending hydrogel for the proliferation of the primary intestinal smooth muscle cells of rats. Biomed Mater, 2013, 8(6): 065002.
4.	李瑞, 王青山. 生物材料生物相容性的評價方法和發展趨勢. 中國組織工程研究, 2011, 15(29): 5471-5474.
5.	Fan MR, Gong M, Da LC, et al. Tissue engineered esophagus scaffold constructed with porcine small intestinal submucosa and synthetic polymers. Biomed Mater, 2014, 9(1): 015012.
6.	D’Amore A, Yoshizumi T, Luketich SK, et al. Bi-layered polyurethane-Extracellular matrix cardiac patch improves ischemic ventricular wall remodeling in a rat model. Biomaterials, 2016, 107: 1-14.
7.	Da LC, Gong M, Chen A, et al. Composite elastomeric polyurethane scaffolds incorporating small intestinal submucosa for soft tissue engineering. Acta Biomater, 2017, 59: 45-57.
8.	李靜, 孫保存, 劉增輝, 等. 2839 例術中冰凍切片診斷與術后石蠟切片診斷的對比研究. 天津醫科大學學報, 2013, 19(6): 471-473.
9.	楊文英. 病理技術 HE 染色在病理診斷中的應用研究. 大家健康 (學術版), 2014, 8(10): 50-51.
10.	王巧. HE 染色方法在臨床病理診斷中的應用. 臨床合理用藥雜志, 2014, 7(32): 118-119.
11.	Ning LJ, Jiang YL, Zhang CH, et al. Fabrication and characterization of a decellularized bovine tendon sheet for tendon reconstruction. J Biomed Mater Res A, 2017, 105(8): 2299-2311.
12.	Huang YZ, Wang JJ, Huang YC, et al. Organic composite-mediated surface coating of human acellular bone matrix with strontium. Mater Sci Eng C Mater Biol Appl, 2018, 84: 12-20.
13.	Xiong BJ, Tan QW, Chen YJ, et al. The effects of platelet-rich plasma and adipose-derived stem cells on neovascularization and fat graft survival. Aesthetic Plast Surg, 2018, 42(1): 1-8.
14.	Yang JL, Yao X, Qing Q, et al. An engineered tendon/ligament bioscaffold derived from decellularized and demineralized cortical bone matrix. J Biomed Mater Res A, 2017, 106(2): 468-478.
15.	Tan QW, Zhang Y, Luo JC, et al. Hydrogel derived from decellularized porcine adipose tissue as a promising biomaterial for soft tissue augmentation. J Biomed Mater Res A, 2017, 105(6): 1756-1764.
16.	晁寧寧, 鄧雪琴, 王瑋, 等. 可注射 CSCl-GP-HEC/SIS 微粒溫敏復合材料軟組織填充初步實驗. 全國組織工程與再生醫學大會, 2013.
17.	吳江鋒, 李紅軍. 制作冰凍切片的體會. 右江民族醫學院學報, 2011, 33(6): 819-820.
18.	胡垚. 富含脂肪及淋巴組織的冰凍切片制作體會. 哈爾濱醫藥, 2015, 35(3): 200-202.
19.	陳敬文, 王天媧, 賴美娜. 碳蠟包埋技術在脂肪染色中的應用研究. 實用醫技雜志, 2016, 23(10): 1133-1134.
20.	王小曉. 水溶性伊紅 Y 不同配置法對 HE 染色效果的影響. 臨床與實驗病理學雜志, 2013, 29(5): 575-576.
21.	張婉儀, 陳哲. 兩種伊紅染色液在 HE 染色中的應用比較. 現代診斷與治療, 2015, 26(12): 2725-2726.
22.	田玉旺, 朱紅艷, 朱培雙, 等. 介紹一種改良的伊紅染色液. 中國組織化學與細胞化學雜志, 2013, 22(4): 328-329.
23.	Wei RQ, Tan B, Tan MY, et al. Grafts of porcine small intestinal submucosa with cultured autologous oral mucosal epithelial cells for esophageal repair in a canine model. Exp Biol Med (Maywood), 2009, 234(4): 453-461.
24.	劉宏銀, 馮喆, 柏林, 等. 雪旺細胞純化培養及體外復合豬小腸黏膜下層的實驗研究. 中國修復重建外科雜志, 2013, 27(6): 743-748.
25.	鄒仲之, 李繼承. 組織學與胚胎學. 8 版. 北京: 人民衛生出版社, 2013: 4.

1. Hu J, Chen B, Guo F, et al. Injectable silk fibroin/polyurethane composite hydrogel for nucleus pulposus replacement. J Mater Sci Mater Med, 2012, 23(3): 711-722.
2. Kuppan P, Sethuraman S, Krishnan UM. PCL and PCL-gelatin nanofibers as esophageal tissue scaffolds: optimization, characterization and cell-matrix interactions. J Biomed Nanotechnology, 2013, 9(9): 1540-1555.
3. Jwo SC, Chin CH, Tang SJ, et al. Tubular scaffolds of gelatin and poly (ε-caprolactone)-block-poly (γ-glutamic acid) blending hydrogel for the proliferation of the primary intestinal smooth muscle cells of rats. Biomed Mater, 2013, 8(6): 065002.
4. 李瑞, 王青山. 生物材料生物相容性的評價方法和發展趨勢. 中國組織工程研究, 2011, 15(29): 5471-5474.
5. Fan MR, Gong M, Da LC, et al. Tissue engineered esophagus scaffold constructed with porcine small intestinal submucosa and synthetic polymers. Biomed Mater, 2014, 9(1): 015012.
6. D’Amore A, Yoshizumi T, Luketich SK, et al. Bi-layered polyurethane-Extracellular matrix cardiac patch improves ischemic ventricular wall remodeling in a rat model. Biomaterials, 2016, 107: 1-14.
7. Da LC, Gong M, Chen A, et al. Composite elastomeric polyurethane scaffolds incorporating small intestinal submucosa for soft tissue engineering. Acta Biomater, 2017, 59: 45-57.
8. 李靜, 孫保存, 劉增輝, 等. 2839 例術中冰凍切片診斷與術后石蠟切片診斷的對比研究. 天津醫科大學學報, 2013, 19(6): 471-473.
9. 楊文英. 病理技術 HE 染色在病理診斷中的應用研究. 大家健康 (學術版), 2014, 8(10): 50-51.
10. 王巧. HE 染色方法在臨床病理診斷中的應用. 臨床合理用藥雜志, 2014, 7(32): 118-119.
11. Ning LJ, Jiang YL, Zhang CH, et al. Fabrication and characterization of a decellularized bovine tendon sheet for tendon reconstruction. J Biomed Mater Res A, 2017, 105(8): 2299-2311.
12. Huang YZ, Wang JJ, Huang YC, et al. Organic composite-mediated surface coating of human acellular bone matrix with strontium. Mater Sci Eng C Mater Biol Appl, 2018, 84: 12-20.
13. Xiong BJ, Tan QW, Chen YJ, et al. The effects of platelet-rich plasma and adipose-derived stem cells on neovascularization and fat graft survival. Aesthetic Plast Surg, 2018, 42(1): 1-8.
14. Yang JL, Yao X, Qing Q, et al. An engineered tendon/ligament bioscaffold derived from decellularized and demineralized cortical bone matrix. J Biomed Mater Res A, 2017, 106(2): 468-478.
15. Tan QW, Zhang Y, Luo JC, et al. Hydrogel derived from decellularized porcine adipose tissue as a promising biomaterial for soft tissue augmentation. J Biomed Mater Res A, 2017, 105(6): 1756-1764.
16. 晁寧寧, 鄧雪琴, 王瑋, 等. 可注射 CSCl-GP-HEC/SIS 微粒溫敏復合材料軟組織填充初步實驗. 全國組織工程與再生醫學大會, 2013.
17. 吳江鋒, 李紅軍. 制作冰凍切片的體會. 右江民族醫學院學報, 2011, 33(6): 819-820.
18. 胡垚. 富含脂肪及淋巴組織的冰凍切片制作體會. 哈爾濱醫藥, 2015, 35(3): 200-202.
19. 陳敬文, 王天媧, 賴美娜. 碳蠟包埋技術在脂肪染色中的應用研究. 實用醫技雜志, 2016, 23(10): 1133-1134.
20. 王小曉. 水溶性伊紅 Y 不同配置法對 HE 染色效果的影響. 臨床與實驗病理學雜志, 2013, 29(5): 575-576.
21. 張婉儀, 陳哲. 兩種伊紅染色液在 HE 染色中的應用比較. 現代診斷與治療, 2015, 26(12): 2725-2726.
22. 田玉旺, 朱紅艷, 朱培雙, 等. 介紹一種改良的伊紅染色液. 中國組織化學與細胞化學雜志, 2013, 22(4): 328-329.
23. Wei RQ, Tan B, Tan MY, et al. Grafts of porcine small intestinal submucosa with cultured autologous oral mucosal epithelial cells for esophageal repair in a canine model. Exp Biol Med (Maywood), 2009, 234(4): 453-461.
24. 劉宏銀, 馮喆, 柏林, 等. 雪旺細胞純化培養及體外復合豬小腸黏膜下層的實驗研究. 中國修復重建外科雜志, 2013, 27(6): 743-748.
25. 鄒仲之, 李繼承. 組織學與胚胎學. 8 版. 北京: 人民衛生出版社, 2013: 4.

Chinese Journal of Reparative and Reconstructive Surgery

A rapid pathological preparation method for composite material observation

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