Native llama Nanobody Library Panning Performed by Phage and Yeast Display Provides Binders Suitable for C-Reactive Protein Detection
C-reactive protein (CRP) is an inflammation biomarker that should be quantified accurately during infections and healing processes. Nanobodies are good candidates to replace conventional antibodies in immunodiagnostics due to their inexpensive production, simple engineering, and the possibility to o...
| Main Authors: | , , , , , , , , , |
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| Format: | Article |
| Language: | English |
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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/2079-6374/11/12/496 |
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| author | Sandra Oloketuyi Robert Bernedo Andreas Christmann Justyna Borkowska Giulia Cazzaniga Horst Wilhelm Schuchmann Joanna Niedziółka-Jönsson Katarzyna Szot-Karpińska Harald Kolmar Ario de Marco |
| author_facet | Sandra Oloketuyi Robert Bernedo Andreas Christmann Justyna Borkowska Giulia Cazzaniga Horst Wilhelm Schuchmann Joanna Niedziółka-Jönsson Katarzyna Szot-Karpińska Harald Kolmar Ario de Marco |
| author_sort | Sandra Oloketuyi |
| collection | DOAJ |
| description | C-reactive protein (CRP) is an inflammation biomarker that should be quantified accurately during infections and healing processes. Nanobodies are good candidates to replace conventional antibodies in immunodiagnostics due to their inexpensive production, simple engineering, and the possibility to obtain higher binder density on capture surfaces. Starting from the same pre-immune library, we compared the selection output resulting from two independent panning strategies, one exclusively exploiting the phage display and another in which a first round of phage display was followed by a second round of yeast display. There was a partial output convergence between the two methods, since two clones were identified using both panning protocols but the first provided several further different sequences, whereas the second favored the recovery of many copies of few clones. The isolated anti-CRP nanobodies had affinity in the low nanomolar range and were suitable for ELISA and immunoprecipitation. One of them was fused to SpyTag and exploited in combination with SpyCatcher as the immunocapture element to quantify CRP using electrochemical impedance spectroscopy. The sensitivity of the biosensor was calculated as low as 0.21 μg/mL. |
| first_indexed | 2024-03-09T04:36:06Z |
| format | Article |
| id | doaj.art-a0d0d89b5196432ebcfb8f266fa9f8c9 |
| institution | Directory Open Access Journal |
| issn | 2079-6374 |
| language | English |
| last_indexed | 2024-03-09T04:36:06Z |
| publishDate | 2021-12-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Biosensors |
| spelling | doaj.art-a0d0d89b5196432ebcfb8f266fa9f8c92023-12-03T13:27:57ZengMDPI AGBiosensors2079-63742021-12-01111249610.3390/bios11120496Native llama Nanobody Library Panning Performed by Phage and Yeast Display Provides Binders Suitable for C-Reactive Protein DetectionSandra Oloketuyi0Robert Bernedo1Andreas Christmann2Justyna Borkowska3Giulia Cazzaniga4Horst Wilhelm Schuchmann5Joanna Niedziółka-Jönsson6Katarzyna Szot-Karpińska7Harald Kolmar8Ario de Marco9Laboratory for Environmental and Life Sciences, University of Nova Gorica, 5000 Nova Gorica, SloveniaLaboratory for Environmental and Life Sciences, University of Nova Gorica, 5000 Nova Gorica, SloveniaApplied Biochemistry, Technical University of Darmstadt, 64200 Darmstadt, GermanyInstitute of Physical Chemistry, Polish Academy of Sciences, 01-224 Warsaw, PolandLaboratory for Environmental and Life Sciences, University of Nova Gorica, 5000 Nova Gorica, SloveniaApplied Biochemistry, Technical University of Darmstadt, 64200 Darmstadt, GermanyInstitute of Physical Chemistry, Polish Academy of Sciences, 01-224 Warsaw, PolandInstitute of Physical Chemistry, Polish Academy of Sciences, 01-224 Warsaw, PolandApplied Biochemistry, Technical University of Darmstadt, 64200 Darmstadt, GermanyLaboratory for Environmental and Life Sciences, University of Nova Gorica, 5000 Nova Gorica, SloveniaC-reactive protein (CRP) is an inflammation biomarker that should be quantified accurately during infections and healing processes. Nanobodies are good candidates to replace conventional antibodies in immunodiagnostics due to their inexpensive production, simple engineering, and the possibility to obtain higher binder density on capture surfaces. Starting from the same pre-immune library, we compared the selection output resulting from two independent panning strategies, one exclusively exploiting the phage display and another in which a first round of phage display was followed by a second round of yeast display. There was a partial output convergence between the two methods, since two clones were identified using both panning protocols but the first provided several further different sequences, whereas the second favored the recovery of many copies of few clones. The isolated anti-CRP nanobodies had affinity in the low nanomolar range and were suitable for ELISA and immunoprecipitation. One of them was fused to SpyTag and exploited in combination with SpyCatcher as the immunocapture element to quantify CRP using electrochemical impedance spectroscopy. The sensitivity of the biosensor was calculated as low as 0.21 μg/mL.https://www.mdpi.com/2079-6374/11/12/496phage displayyeast displaynanobodiesbiopanningCRP |
| spellingShingle | Sandra Oloketuyi Robert Bernedo Andreas Christmann Justyna Borkowska Giulia Cazzaniga Horst Wilhelm Schuchmann Joanna Niedziółka-Jönsson Katarzyna Szot-Karpińska Harald Kolmar Ario de Marco Native llama Nanobody Library Panning Performed by Phage and Yeast Display Provides Binders Suitable for C-Reactive Protein Detection Biosensors phage display yeast display nanobodies biopanning CRP |
| title | Native llama Nanobody Library Panning Performed by Phage and Yeast Display Provides Binders Suitable for C-Reactive Protein Detection |
| title_full | Native llama Nanobody Library Panning Performed by Phage and Yeast Display Provides Binders Suitable for C-Reactive Protein Detection |
| title_fullStr | Native llama Nanobody Library Panning Performed by Phage and Yeast Display Provides Binders Suitable for C-Reactive Protein Detection |
| title_full_unstemmed | Native llama Nanobody Library Panning Performed by Phage and Yeast Display Provides Binders Suitable for C-Reactive Protein Detection |
| title_short | Native llama Nanobody Library Panning Performed by Phage and Yeast Display Provides Binders Suitable for C-Reactive Protein Detection |
| title_sort | native llama nanobody library panning performed by phage and yeast display provides binders suitable for c reactive protein detection |
| topic | phage display yeast display nanobodies biopanning CRP |
| url | https://www.mdpi.com/2079-6374/11/12/496 |
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