title: Evaluation of Decalcification Techniques for Rat Femurs Using HE and Immunohistochemical Staining.
abstract: Aim. In routine histopathology, decalcification is an essential step for mineralized tissues. The purpose of this study is to evaluate the effects of different decalcification solutions on the morphological and antigenicity preservation in Sprague Dawley (SD) rat femurs.Materials and Methods. Four different decalcification solutions were employed to remove the mineral substances from rat femurs, including 10% neutral buffered EDTA, 3% nitric acid, 5% nitric acid, and 8% hydrochloric acid/formic acid. Shaking and low temperature were used to process the samples. The stainings of hematoxylin-eosin (HE) and immunohistochemical (IHC) were employed to evaluate the bone morphology and antigenicity.Key Findings. Different decalcification solutions may affect the quality of morphology and the staining of paraffin-embedded sections in pathological examinations. Among four decalcifying solutions, 3% nitric acid is the best decalcifying agent for HE staining. 10% neutral buffered EDTA and 5% nitric acid are the preferred decalcifying agents for IHC staining.Significance. The current study investigated the effects of different decalcifying agents on the preservation of the bone structure and antigenicity, which will help to develop suitable protocols for the analyses of the bony tissue.
authors.affiliation: Preclinical Medicine School, Beijing University of Chinese Medicine, Beijing 100029, China.
authors.forename: Haixia
authors.initials: H
authors.lastname: Liu
authors.affiliation: Preclinical Medicine School, Beijing University of Chinese Medicine, Beijing 100029, China.
authors.forename: Ruyuan
authors.initials: R
authors.lastname: Zhu
authors.affiliation: Chinese Material Medica School, Beijing University of Chinese Medicine, Beijing 100029, China.
authors.forename: Chenyue
authors.initials: C
authors.lastname: Liu
authors.affiliation: Preclinical Medicine School, Beijing University of Chinese Medicine, Beijing 100029, China.
authors.forename: Rufeng
authors.initials: R
authors.lastname: Ma
authors.affiliation: Preclinical Medicine School, Beijing University of Chinese Medicine, Beijing 100029, China.
authors.forename: Lili
authors.initials: L
authors.lastname: Wang
authors.affiliation: Preclinical Medicine School, Beijing University of Chinese Medicine, Beijing 100029, China.
authors.forename: Beibei
authors.initials: B
authors.lastname: Chen
authors.affiliation: Preclinical Medicine School, Beijing University of Chinese Medicine, Beijing 100029, China.
authors.forename: Lin
authors.initials: L
authors.lastname: Li
authors.affiliation: Preclinical Medicine School, Beijing University of Chinese Medicine, Beijing 100029, China.
authors.forename: Jianzhao
authors.initials: J
authors.lastname: Niu
authors.affiliation: Diabetes Research Center, Beijing University of Chinese Medicine, Beijing 100029, China.
authors.forename: Dandan
authors.identifier: 0000-0002-0757-984X
authors.initials: D
authors.lastname: Zhao
authors.affiliation: Diabetes Research Center, Beijing University of Chinese Medicine, Beijing 100029, China.
authors.forename: Fangfang
authors.initials: F
authors.lastname: Mo
authors.affiliation: The Research Institute of McGill University Health Center, Montreal, QC, Canada H4A 3J1.
authors.forename: Min
authors.initials: M
authors.lastname: Fu
authors.affiliation: Oral Biological Medicinal Science, University of British Columbia, Vancouver, BC, Canada V6T 1Z3.
authors.forename: Dieter
authors.initials: D
authors.lastname: Brömme
authors.affiliation: Diabetes Research Center, Beijing University of Chinese Medicine, Beijing 100029, China.
authors.forename: Dongwei
authors.identifier: 0000-0002-7793-1482
authors.initials: D
authors.lastname: Zhang
authors.affiliation: Diabetes Research Center, Beijing University of Chinese Medicine, Beijing 100029, China.
authors.forename: Sihua
authors.identifier: 0000-0002-0332-3767
authors.initials: S
authors.lastname: Gao
mesh_terms: D000818:Animals; D000941:Antigens / Q000378:metabolism; D003648:Decalcification Technique / Q000379:methods*; D004801:Eosine Yellowish-(YS) / Q000378:metabolism*; D005260:Female; D005269:Femur / Q000473:pathology*; D006416:Hematoxylin / Q000378:metabolism*; D007091:Image Processing, Computer-Assisted; D007150:Immunohistochemistry / Q000379:methods*; D017207:Rats, Sprague-Dawley; D012996:Solutions; D013194:Staining and Labeling; D013997:Time Factors
chemical_list: D000941:Antigens; D012996:Solutions; D004801:Eosine Yellowish-(YS); D006416:Hematoxylin