期刊
JOURNAL OF HISTOCHEMISTRY & CYTOCHEMISTRY
卷 64, 期 1, 页码 18-31出版社
SAGE PUBLICATIONS LTD
DOI: 10.1369/0022155415616162
关键词
sugars; disaccharide; lactose; sucrose; antigen masking; antibody; multiple immunostaining; elution; stripping; freeze-drying
类别
资金
- GlaxoSmithKline
- Azienda Ospedaliera San Gerardo [HGS1006-C1121]
- Fondazione per la Ricerca Scientifica Termale (FoRST)
- IV call grants (Project Lymphopoiesis In Secondary Lymphoid Tissue'')
- Regione Lombardia [DDG 6716]
- Departmental University of Milano-Bicocca
- Hospital funds
Drying of the tissue section, partial or total, during immunostaining negatively affects both the staining of tissue antigens and the ability to remove previously deposited antibody layers, particularly during sequential rounds of de-staining and re-staining for multiple antigens. The cause is a progressive loss of the protein-associated water up to the removal of the non-freezable water, a step which abolishes the immunoavailability of the epitope. In order to describe and prevent these adverse effects, we tested, among other substances, sugars, which are known to protect unicellular organisms from freezing and dehydration, and stabilize drugs and reagents in solid state form in medical devices. Disaccharides (lactose, sucrose) prevented the air drying-induced antigen masking and protected tissue-bound antigens and antibodies from air drying-induced damage. Complete removal of the bound antibody layers by chemical stripping was permitted if lactose was present during air drying. Lactose, sucrose and other disaccharides prevent air drying artifacts, allow homogeneous, consistent staining and the reuse of formalin-fixed, paraffin-embedded tissue sections for repeated immunostaining rounds by guaranteeing constant staining quality in suboptimal hydration conditions.
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