Efficient small-scale conjugation of DNA to primary antibodies for multiplexed cellular targeting
| العنوان: | Efficient small-scale conjugation of DNA to primary antibodies for multiplexed cellular targeting |
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| المؤلفون: | Tom F. A. de Greef, Roger Riera Brillas, Lorenzo Albertazzi, Glenn A.O. Cremers, Bas J.H.M. Rosier |
| المساهمون: | Chemical Biology, Computational Biology, Molecular Biosensing for Med. Diagnostics |
| المصدر: | Bioconjugate Chemistry, 30, 9-10, pp. 2384-2392 Bioconjugate Chemistry, 30(9), 2384-2392. American Chemical Society bioRxiv Bioconjugate Chemistry Bioconjugate Chem. Bioconjugate Chemistry, 30, 2384-2392 |
| بيانات النشر: | American Chemical Society, 2019. |
| سنة النشر: | 2019 |
| مصطلحات موضوعية: | Models, Molecular, Protein Conformation, Biomedical Engineering, Pharmaceutical Science, Bioengineering, 02 engineering and technology, Computational biology, 01 natural sciences, Antibodies, Article, 03 medical and health sciences, chemistry.chemical_compound, Affinity Reagent, Bacterial Proteins, DNA nanotechnology, Animals, Humans, Bovine serum albumin, 030304 developmental biology, Pharmacology, 0303 health sciences, biology, 010405 organic chemistry, Oligonucleotide, Organic Chemistry, DNA, 021001 nanoscience & nanotechnology, Primary and secondary antibodies, 0104 chemical sciences, Oligodeoxyribonucleotides, chemistry, biology.protein, Protein G, Biophysical Chemistry, 0210 nano-technology, Biotechnology, Conjugate |
| الوصف: | The combination of the specificity of antibodies and the programmability of DNA nanotechnology has provided the scientific community with a powerful tool to label and unambiguously distinguish a large number of subcellular targets using fluorescence-based read-out methods. While primary antibodies are commercially available for a large class of targets, a general stoichiometric site-specific DNA labeling strategy for this affinity reagent is lacking. Here, we present a universal, site-selective, conjugation method using a small photocrosslinkable protein G adaptor that allows labeling of antibodies of different host species with a controlled number of short oligonucleotides (ODNs). Importantly, we illustrate that this conjugation method can be directly performed on commercially-available primary antibodies, on a small scale and without cross-reactivity towards other proteins, such as bovine serum albumin. In addition, we present a general, benchtop-compatible strategy to purify DNA-labeled antibodies without loss of function. The application of protein G-ODN labeled primary antibodies is demonstrated by employing three well-known methods for detecting subcellular targets using fluorescent read-out, including flow cytometry, DNA-PAINT, and dSTORM. This work thus establishes a general and efficient platform for the synthesis of a library of unique ODN-antibody conjugates, facilitating the broader use of DNA-based programmable tags for multiplexed labeling to identify subcellular features with nanometer-precision, improving our understanding of cellular structure and function. |
| وصف الملف: | application/pdf |
| اللغة: | English |
| تدمد: | 1520-4812 1043-1802 |
| URL الوصول: | https://explore.openaire.eu/search/publication?articleId=doi_dedup___::971738277279cf390843c9fe4da0f1c0 https://research.tue.nl/en/publications/af986012-0008-40c4-8a8a-731d71f28125 |
| حقوق: | OPEN |
| رقم الأكسشن: | edsair.doi.dedup.....971738277279cf390843c9fe4da0f1c0 |
| قاعدة البيانات: | OpenAIRE |
Find this issue from the American Chemical Society | تدمد: | 15204812 10431802 |
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