Immunocytochemical staining of endogenous nuclear proteins with the HIS-1 anti-poly-histidine monoclonal antibody: A potential source of error in His-tagged protein detection

Article


Chilumuri, A., Markiv, Anatoliy and Milton, Nathaniel G.N. 2014. Immunocytochemical staining of endogenous nuclear proteins with the HIS-1 anti-poly-histidine monoclonal antibody: A potential source of error in His-tagged protein detection. Acta Histochemica. 116 (6), pp. 1022-1028.
AuthorsChilumuri, A., Markiv, Anatoliy and Milton, Nathaniel G.N.
Abstract

Histidine-tagged proteins are widely used in biochemical studies and frequently detected with antibodies specific for the histidine tag. Immunocytochemistry is widely used in studies with overexpressed proteins to determine cellular localization and in the case of histidine-tagged proteins can be carried out with anti-polyhistidine antibodies. Recent studies have suggested that polyhistidine sequences are present within a small number of human proteins and may direct expression to the nucleus and nuclear speckles compartments of the cell. In this study immunocytochemical staining of human SH-SY5Y neuroblastoma cell lines with the HIS-1 anti-polyhistidine monoclonal antibody were determined. Results showed that the HIS-1 anti-polyhistidine monoclonal antibody stained endogenous nuclear proteins in SH-SY5Y cells. The stained proteins were contained within the nuclear membrane, but were not directly linked to DNA. In a histidine-tagged catalase overexpressing cell line the HIS-1 anti-polyhistidine monoclonal antibody showed nuclear staining, whilst staining with the CAT-505 anti-catalase monoclonal antibody showed primarily cytoplasmic staining. These results suggest that anti-polyhistidine antibody staining shows significant cross-reactivity with endogenous nuclear proteins in SH-SY5Y neuroblastoma cells and may not be suitable for localization studies of histidine-tagged proteins. Immunocytochemical studies with anti-polyhistidine antibodies and localization of histidine-tagged proteins must be confirmed with protein specific antibodies or other methodology.

KeywordsImmunocytochemistry; SH-SY5Y neuroblastoma; Anti-polyhistidine; Transfection; His-tagged proteins; Catalase overexpression
JournalActa Histochemica
Journal citation116 (6), pp. 1022-1028
ISSN0065-1281
1618-0372
Year2014
PublisherElsevier for Urban and Fischer
Digital Object Identifier (DOI)doi:10.1016/j.acthis.2014.04.006
Publication dates
Print14 May 2014
Publication process dates
Deposited03 Jul 2017
Accepted08 Apr 2014
FunderUniversity of Westminster
WestFocus
UK Department of Trade and Industry
University of Westminster
Department of Trade and Industry
WestFocus
Copyright information© 2014 Elsevier
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