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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| Format: | Article |
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MDPI AG
2021-12-01
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| Series: | Biosensors |
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| Online Access: | https://www.mdpi.com/2079-6374/11/12/524 |
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| author | Tania García-Maceira Fé I. García-Maceira José A. González-Reyes Luis A. Torres-Sánchez Ana Belén Aragón-Gómez María Eugenia García-Rubiño Elier Paz-Rojas |
| author_facet | Tania García-Maceira Fé I. García-Maceira José A. González-Reyes Luis A. Torres-Sánchez Ana Belén Aragón-Gómez María Eugenia García-Rubiño Elier Paz-Rojas |
| author_sort | Tania García-Maceira |
| collection | DOAJ |
| 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. |
| first_indexed | 2024-03-09T04:35:41Z |
| format | Article |
| id | doaj.art-38bdac45845e4ee7abcdbf022f14e60a |
| institution | Directory Open Access Journal |
| issn | 2079-6374 |
| language | English |
| last_indexed | 2024-03-09T04:35:41Z |
| publishDate | 2021-12-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Biosensors |
| spelling | doaj.art-38bdac45845e4ee7abcdbf022f14e60a2023-12-03T13:28:27ZengMDPI AGBiosensors2079-63742021-12-01111252410.3390/bios11120524Covalent Immobilization of Antibodies through Tetrazine-TCO Reaction to Improve Sensitivity of ELISA TechniqueTania García-Maceira0Fé I. García-Maceira1José A. González-Reyes2Luis A. Torres-Sánchez3Ana Belén Aragón-Gómez4María Eugenia García-Rubiño5Elier Paz-Rojas6Canvax Biotech, Parque Científico y Tecnológico Rabanales 21, c/Astrónoma Cecilia Payne s/n, Edificio Orión, 14014 Córdoba, SpainCanvax Biotech, Parque Científico y Tecnológico Rabanales 21, c/Astrónoma Cecilia Payne s/n, Edificio Orión, 14014 Córdoba, SpainDepartment of Cell Biology, Physiology and Immunology, Campus de Excelencia Internacional Agroalimentario (ceiA3), University of Córdoba, 14014 Córdoba, SpainCanvax Biotech, Parque Científico y Tecnológico Rabanales 21, c/Astrónoma Cecilia Payne s/n, Edificio Orión, 14014 Córdoba, SpainCanvax Biotech, Parque Científico y Tecnológico Rabanales 21, c/Astrónoma Cecilia Payne s/n, Edificio Orión, 14014 Córdoba, SpainDepartment of Physical Chemistry, Faculty of Pharmacy, University of Granada, 18071 Granada, SpainCanvax Biotech, Parque Científico y Tecnológico Rabanales 21, c/Astrónoma Cecilia Payne s/n, Edificio Orión, 14014 Córdoba, SpainEnzyme-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.https://www.mdpi.com/2079-6374/11/12/524ELISAantibodies immobilizationtetrazineTCOCEA |
| spellingShingle | Tania García-Maceira Fé I. García-Maceira José A. González-Reyes Luis A. Torres-Sánchez Ana Belén Aragón-Gómez María Eugenia García-Rubiño Elier Paz-Rojas Covalent Immobilization of Antibodies through Tetrazine-TCO Reaction to Improve Sensitivity of ELISA Technique Biosensors ELISA antibodies immobilization tetrazine TCO CEA |
| 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 | ELISA antibodies immobilization tetrazine TCO CEA |
| url | https://www.mdpi.com/2079-6374/11/12/524 |
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