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Proceedings Paper

Oriented conjugates of monoclonal and single-domain antibodies with quantum dots for flow cytometry and immunohistochemistry diagnostic applications
Author(s): Alyona Sukhanova; Klervi Even-Desrumeaux; Jean-Marc Millot; Patrick Chames; Daniel Baty; Mikhail Artemyev; Vladimir Oleinikov; Jacques H. M. Cohen; Igor Nabiev
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Paper Abstract

Ideal diagnostic nanoprobes should not exceed 15 nm in size and should contain high-affinity homogeneously oriented capture molecules on their surface. An advanced procedure for antibody (Ab) reduction was used to cleave each Ab into two functional half-Abs, 75-kDa heavy-light chain fragments, each containing an intact antigen-binding site. Affinity purification of half-Abs followed by their linkage to quantum dots (QDs) yielded oriented QD-Ab conjugates whose functionality was considerably improved compared to those obtained using the standard protocols. Ultrasmall diagnostic nanoprobes were engineered through oriented conjugation of QDs with 13-kDa single-domain Abs (sdAbs) derived from llama IgG. sdAbs were tagged with QDs via an additional cysteine residue specifically integrated into the C-terminal region of sdAb using genetic engineering. This approach made it possible to obtain sdAb-QD nanoprobes <12 nm in diameter comprising four copies of sdAbs linked to the same QD in an oriented manner. sdAb-QD conjugates against carcinoembryonic antigen (CEA) and HER2 exhibited an extremely high specificity in flow cytometry; the quality of immunohistochemical labeling of biopsy samples was found to be superior to that of labeling according to the current "gold standard" protocols of anatomo-pathological practice. The nano-bioengineering approaches developed can be extended to oriented conjugation of Abs and sdAbs with different semiconductor, noble metal, or magnetic nanoparticles.

Paper Details

Date Published: 2 February 2012
PDF: 9 pages
Proc. SPIE 8232, Colloidal Nanocrystals for Biomedical Applications VII, 82320T (2 February 2012); doi: 10.1117/12.905896
Show Author Affiliations
Alyona Sukhanova, Moscow Engineering Physics Institute (Russian Federation)
Univ. de Reims Champagne-Ardenne (France)
Klervi Even-Desrumeaux, INSERM (France)
Jean-Marc Millot, Univ. de Reims Champagne-Ardenne (France)
Patrick Chames, INSERM (France)
Daniel Baty, INSERM (France)
Mikhail Artemyev, Moscow Engineering Physics Institute (Russian Federation)
Belarusian State Univ. (Belarus)
Vladimir Oleinikov, Moscow Engineering Physics Institute (Russian Federation)
Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry (Russian Federation)
Jacques H. M. Cohen, Univ. de Reims Champagne-Ardenne (France)
Igor Nabiev, Moscow Engineering Physics Institute (Russian Federation)
Univ. de Reims Champagne-Ardenne (France)

Published in SPIE Proceedings Vol. 8232:
Colloidal Nanocrystals for Biomedical Applications VII
Wolfgang J. Parak; Kenji Yamamoto; Marek Osinski, Editor(s)

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