A novel methodology for double protein A-gold immunolabelling utilizing the monovalent fragment of protein A

Thorpe, Julian (1992) A novel methodology for double protein A-gold immunolabelling utilizing the monovalent fragment of protein A. Journal of Histochemistry and Cytochemistry, 40 (3). pp. 435-441. ISSN 0022-1554

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Abstract

A method for sequential protein A-gold immunolabeling is described whereby the binding of second gold probe to the first antibody-protein A-gold complex is reduced to acceptably minimal levels. Immunolabeling of thin sections of embedded pituitary tissue was used as a model system. After an initial immunolabeling for prolactin, sections were incubated in normal serum (rabbit) followed by a monovalent fragment of protein A. These latter two incubations reduced artifactual second gold probe label over prolactin-labeled secretory granules to minimal levels (much less than 1 particle per granule) when sections were subsequently immunolabeled with normal serum. The combination of normal serum and protein A fragment incubations saturates IgG and protein A binding sites on the first antibody-gold probe complex. The latter is thereafter unable to bind further IgG (and thus gold probe) because of the monovalent nature of the protein A fragment. It is suggested that this methodology may be extended to multiple immunolabeling procedures for electron microscopy. In addition, when used before single labeling this method may be an effective way to minimize nonspecific IgG binding in cases where the tissue or antibody under study may be a problem.

Item Type: Article
Schools and Departments: School of Life Sciences > Biochemistry
Depositing User: Julian Thorpe
Date Deposited: 06 Feb 2012 18:40
Last Modified: 16 Jul 2012 09:40
URI: http://sro.sussex.ac.uk/id/eprint/17686
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