Simultaneous detection of multiple targets for ultrastructural immunocytochemistry

0434293 - ÚMG 2015 RIV DE eng J - Journal Article
Philimonenko, Vlada - Filimonenko, Anatolij - Šloufová, I. - Hrubý, Martin - Novotný, F. - Halbhuber, Z. - Krivjanská, M. - Nebesářová, Jana - Šlouf, Miroslav - Hozák, Pavel
Simultaneous detection of multiple targets for ultrastructural immunocytochemistry.
Histochemistry and Cell Biology. Roč. 141, č. 3 (2014), s. 229-239. ISSN 0948-6143. E-ISSN 1432-119X
R&D Projects: GA AV ČR KAN200520704; GA TA ČR TE01020118; GA ČR GAP305/11/2232; GA MŠMT LD12063
Grant - others:Human Frontier Science Program(FR) RGP0017/2013
Institutional research plan: CEZ:AV0Z50520514
Institutional support: RVO:61389013 ; RVO:60077344 ; RVO:68378050
Keywords : Immunolabeling * Metal nanoparticles * Electron microscopy * Cell nucleus * Ultrastructure * Phosphatidylinositol-4,5-Bisphosphate (PIP2)
Subject RIV: EB - Genetics ; Molecular Biology
Impact factor: 3.054, year: 2014

Simultaneous detection of biological molecules by means of indirect immunolabeling provides valuable information about their localization in cellular compartments and their possible interactions in macromolecular complexes. While fluorescent microscopy allows for simultaneous detection of multiple antigens, the sensitive electron microscopy immunodetection is limited to only two antigens. In order to overcome this limitation, we prepared a set of novel, shape-coded metal nanoparticles readily discernible in transmission electron microscopy which can be conjugated to antibodies or other bioreactive molecules. With the use of novel nanoparticles, various combinations with commercial gold nanoparticles can be made to obtain a set for simultaneous labeling. For the first time in ultrastructural histochemistry, up to five molecular targets can be identified simultaneously. We demonstrate the usefulness of the method by mapping of the localization of nuclear lipid phosphatidylinositol-4,5-bisphosphate together with four other molecules crucial for genome function, which proves its suitability for a wide range of biomedical applications.
Permanent Link: http://hdl.handle.net/11104/0241064