The contribution of bifunctional SkipDewax pretreatment solution, rabbit monoclonal antibodies, and polymer detection systems in immunohistochemistry.
BACKGROUND
In immunohistochemistry, nonstandardized antigen retrieval protocols and fluids, poor-quality antibodies, and the presence of endogenous biotin frequently lead to incorrect results. Recently, advanced reagents including bifunctional SkipDewax pretreatment solution (BSPS), rabbit monoclonal (RM) antibodies, and biotin-free polymer detection systems (PDSs) have been developed, which, it is claimed, resolve these problems.


OBJECTIVE
To determine whether BSPS, RM antibodies, and biotin-free PDSs improve the accuracy of immunohistochemistry; to optimize a new protocol consisting of a combination of BSPS, RM antibodies, and PDSs; and to compare it with a conventional protocol.


METHODS
The efficacies of BSPS, RM antibodies, and PDSs were compared with those of their respective conventional reagents using multitissue spring-roll sections. The new protocol was compared with a conventional protocol using Ki-67 immunostaining of 49 colorectal carcinoma specimens.


RESULTS
For antigen retrieval, BSPS resulted in similar or better tissue staining than an EDTA solution, but the efficacy of BSPS decreased when it was reused. Most RM antibodies resulted in a greater proportion of positive cells than the corresponding non-RM antibodies, which did not produce satisfactory results in the absence of antigen retrieval. The PDSs Bond, ChemMate, and SuperPicture resulted in a high percentage of positive cells, good staining intensities, and low backgrounds. Other PDSs, except that from Ventana, resulted in high backgrounds and false positivity. The new combined protocol resulted in better Ki-67 staining than the conventional assay.


CONCLUSIONS
Bifunctional SkipDewax pretreatment solution, RM antibodies, and PDSs improve staining quality and diagnostic accuracy of immunohistochemistry assays and provide a foundation for standardization.
Wong
Sze Chuen Cesar
SC
Department of Clinical Oncology at the Sir Y. K. Pao Centre for Cancer, The Chinese University of Hong Kong, Prince of Wales Hospital, Hong Kong Special Administrative Region, China. cesar01@netvigator.com
Chan
John K C
JK
Lo
Elena S F
ES
Chan
Amanda K C
AK
Wong
Manson C K
MC
Chan
Charles M L
CM
Lam
Money Y Y
MY
Chan
Anthony T C
AT
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D001430:Bacteriocins; D017252:CD3 Complex / Q000032:analysis / Q000276:immunology; D018956:CD5 Antigens / Q000032:analysis / Q000276:immunology; D015179:Colorectal Neoplasms / Q000737:chemistry*; D003432:Cross-Linking Reagents; D019938:Cyclin D1 / Q000032:analysis / Q000276:immunology; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods* / Q000592:standards; D019394:Ki-67 Antigen / Q000032:analysis* / Q000276:immunology; D011108:Polymers; D011817:Rabbits; D011933:Reagent Kits, Diagnostic; D018719:Erb-b2 Receptor Tyrosine Kinases / Q000032:analysis / Q000276:immunology; D013194:Staining and Labeling; D016708:Synaptophysin / Q000032:analysis / Q000276:immunology
D000911:Antibodies, Monoclonal; D001430:Bacteriocins; D017252:CD3 Complex; D018956:CD5 Antigens; D003432:Cross-Linking Reagents; D019938:Cyclin D1; D019394:Ki-67 Antigen; D011108:Polymers; D011933:Reagent Kits, Diagnostic; D018719:Erb-b2 Receptor Tyrosine Kinases; D016708:Synaptophysin; C100883:dextranicin-24