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An effective and practical immunohistochemical protocol for bone specimens characterized by hyaluronidase and pepsin predigestion combined with alkaline phosphatase-mediated chromogenic detection

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dc.contributor.authorLi, Shangfu
dc.contributor.authorLiu, Bin
dc.contributor.authorTian, Ming
dc.contributor.authorZhang, Liangming
dc.contributor.authorTickner, Jennifer
dc.contributor.authorXu, Jiake
dc.contributor.authorRong, Limin
dc.date.accessioned2020-03-18T09:25:09Z
dc.date.available2020-03-18T09:25:09Z
dc.date.issued2015
dc.description.abstractThe aim of this study was to provide an effective procedure for immunohistochemistry (IHC) investigations of bone specimens. Samples from rat femoral and human vertebral bone were processed with a detailed and effective IHC protocol summarized here. First, a novel antigen retrieval (AR) method of hyaluronidase combined pepsin predigestion (H+P) was established and the optimal concentration and pH value for AR of bone specimens were determined. Second, the newly developed method was compared with existing AR methods (boiling in sodium citrate, hyaluronidase predigestion (H) and pepsin predigestion (P), with PBS only as the negative control) using two chromogenic detection systems (horseradish peroxidase (HRP) and alkaline phosphatase (AP)) to evaluate their efficacy in obtaining the best IHC results for bone samples. Considering the drawbacks of significant shrinking and detachment from slide for heat retrieval methods and the only moderate immunolabeling for H and P, H+P was the optimal AR method for IHC of bone specimens with the advantages of both good morphological preservation and strong immunoreactivity. Moreover, AP-mediated chromogenic detection was superior to HRP-labeled chromogenic detection due to significantly less nonspecific staining. In conclusion, we presented an effective and practical IHC protocol for bone specimens characterized by H+P predigestion combined with APmediated chromogenic detection. Finally, a detailed troubleshooting guide was provided for common mistakes that occur during IHC processing of the bone tissue samples.es
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dc.format.extent13es
dc.identifier.citationHistology and Histopathology, Vol. 30, n.º 3 (2015)
dc.identifier.doihttps://doi.org/10.14670/HH-30.331
dc.identifier.issn1699-5848
dc.identifier.issn0213-3911
dc.identifier.urihttp://hdl.handle.net/10201/88201
dc.languageenges
dc.publisherF. Hernández y Juan F. Madrid. Universidad de Murcia: Departamento de Biología Celular e Histologíaes
dc.rightsinfo:eu-repo/semantics/openAccesses
dc.rightsAttribution-NonCommercial-NoDerivatives 4.0 International*
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/4.0/*
dc.subjectImmunohistochemistryes
dc.subjectBone tissuees
dc.subjectAntigen retrievales
dc.subjectHyaluronidasees
dc.subject.otherCDU::5 - Ciencias puras y naturales::57 - Biología::576 - Biología celular y subcelular. Citologíaes
dc.titleAn effective and practical immunohistochemical protocol for bone specimens characterized by hyaluronidase and pepsin predigestion combined with alkaline phosphatase-mediated chromogenic detectiones
dc.typeinfo:eu-repo/semantics/articlees
dspace.entity.typePublicationes
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Histology and histopathology Vol.30, nº3 (2015)