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Covalent Immobilization of Antibodies through Tetrazine-TCO Reaction to Improve Sensitivity of ELISA Technique

Enzyme-linked immunosorbent assay (ELISA) is routinely used to detect biomolecules related to several diseases facilitating diagnosis and monitoring of these, as well as the possibility of decreasing their mortality rate. Several methods have been carried out to improve the ELISA sensitivity through...

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Autores principales: García-Maceira, Tania, García-Maceira, Fé I., González-Reyes, José A., Torres-Sánchez, Luis A., Aragón-Gómez, Ana Belén, García-Rubiño, María Eugenia, Paz-Rojas, Elier
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8699711/
https://www.ncbi.nlm.nih.gov/pubmed/34940281
http://dx.doi.org/10.3390/bios11120524
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author García-Maceira, Tania
García-Maceira, Fé I.
González-Reyes, José A.
Torres-Sánchez, Luis A.
Aragón-Gómez, Ana Belén
García-Rubiño, María Eugenia
Paz-Rojas, Elier
author_facet García-Maceira, Tania
García-Maceira, Fé I.
González-Reyes, José A.
Torres-Sánchez, Luis A.
Aragón-Gómez, Ana Belén
García-Rubiño, María Eugenia
Paz-Rojas, Elier
author_sort García-Maceira, Tania
collection PubMed
description Enzyme-linked immunosorbent assay (ELISA) is routinely used to detect biomolecules related to several diseases facilitating diagnosis and monitoring of these, as well as the possibility of decreasing their mortality rate. Several methods have been carried out to improve the ELISA sensitivity through antibodies immobilization on the microtiter plates. Here, we have developed a strategy of antibodies immobilization to improve the ELISA sensitivity increasing the antibody density surface through the tetrazine (Tz)-trans-cyclooctene (TCO) reaction. For this, we prepared surfaces with tetrazine groups while the captured antibody was conjugated with TCO. The tetrazine surfaces were prepared in two different ways: (1) from aminated plates and (2) from Tz-BSA-coated plates. The surfaces were evaluated using two sandwich ELISA models, one of them using the low-affinity antibody anti-c-myc as a capture antibody to detect the c-myc-GST-IL8h recombinant protein, and the other one to detect the carcinoembryonic human protein (CEA). The sensitivity increased in both surfaces treated with tetrazine in comparison with the standard unmodified surface. The c-myc-GST-IL8h detection was around 10-fold more sensible on both tetrazine surfaces, while CEA ELISA detection increased 12-fold on surfaces coated with Tz-BSA. In conclusion, we show that it is possible to improve the ELISA sensitivity using this immobilization system, where capture antibodies bond covalently to surfaces.
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spelling pubmed-86997112021-12-24 Covalent Immobilization of Antibodies through Tetrazine-TCO Reaction to Improve Sensitivity of ELISA Technique García-Maceira, Tania García-Maceira, Fé I. González-Reyes, José A. Torres-Sánchez, Luis A. Aragón-Gómez, Ana Belén García-Rubiño, María Eugenia Paz-Rojas, Elier Biosensors (Basel) Article Enzyme-linked immunosorbent assay (ELISA) is routinely used to detect biomolecules related to several diseases facilitating diagnosis and monitoring of these, as well as the possibility of decreasing their mortality rate. Several methods have been carried out to improve the ELISA sensitivity through antibodies immobilization on the microtiter plates. Here, we have developed a strategy of antibodies immobilization to improve the ELISA sensitivity increasing the antibody density surface through the tetrazine (Tz)-trans-cyclooctene (TCO) reaction. For this, we prepared surfaces with tetrazine groups while the captured antibody was conjugated with TCO. The tetrazine surfaces were prepared in two different ways: (1) from aminated plates and (2) from Tz-BSA-coated plates. The surfaces were evaluated using two sandwich ELISA models, one of them using the low-affinity antibody anti-c-myc as a capture antibody to detect the c-myc-GST-IL8h recombinant protein, and the other one to detect the carcinoembryonic human protein (CEA). The sensitivity increased in both surfaces treated with tetrazine in comparison with the standard unmodified surface. The c-myc-GST-IL8h detection was around 10-fold more sensible on both tetrazine surfaces, while CEA ELISA detection increased 12-fold on surfaces coated with Tz-BSA. In conclusion, we show that it is possible to improve the ELISA sensitivity using this immobilization system, where capture antibodies bond covalently to surfaces. MDPI 2021-12-20 /pmc/articles/PMC8699711/ /pubmed/34940281 http://dx.doi.org/10.3390/bios11120524 Text en © 2021 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
García-Maceira, Tania
García-Maceira, Fé I.
González-Reyes, José A.
Torres-Sánchez, Luis A.
Aragón-Gómez, Ana Belén
García-Rubiño, María Eugenia
Paz-Rojas, Elier
Covalent Immobilization of Antibodies through Tetrazine-TCO Reaction to Improve Sensitivity of ELISA Technique
title Covalent Immobilization of Antibodies through Tetrazine-TCO Reaction to Improve Sensitivity of ELISA Technique
title_full Covalent Immobilization of Antibodies through Tetrazine-TCO Reaction to Improve Sensitivity of ELISA Technique
title_fullStr Covalent Immobilization of Antibodies through Tetrazine-TCO Reaction to Improve Sensitivity of ELISA Technique
title_full_unstemmed Covalent Immobilization of Antibodies through Tetrazine-TCO Reaction to Improve Sensitivity of ELISA Technique
title_short Covalent Immobilization of Antibodies through Tetrazine-TCO Reaction to Improve Sensitivity of ELISA Technique
title_sort covalent immobilization of antibodies through tetrazine-tco reaction to improve sensitivity of elisa technique
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8699711/
https://www.ncbi.nlm.nih.gov/pubmed/34940281
http://dx.doi.org/10.3390/bios11120524
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