Genetic fusion of single-chain variable fragments to partial spider silk improves target detection in micro- and nanoarrays.
(2015) In Biotechnology Journal 11(3). p.437-448- Abstract
- Immobilizing biomolecules with retained functionality and stability on solid supports is crucial for generation of sensitive immunoassays. However, upon use of conventional immobilization strategies, a major portion of the biomolecules (e.g. antibodies) frequently tends to lose their bioactivity. In this study, we describe a procedure to immobilize human single-chain variable fragment (scFv) via genetic fusion to partial spider silk, which have a high tendency to adhere to solid supports. Two scFvs, directed towards serum proteins, were genetically fused to partial spider silk proteins and expressed as silk fusion proteins in E. coli. Antigen binding ability of scFvs attached to a partial silk protein denoted RC was investigated using... (More)
- Immobilizing biomolecules with retained functionality and stability on solid supports is crucial for generation of sensitive immunoassays. However, upon use of conventional immobilization strategies, a major portion of the biomolecules (e.g. antibodies) frequently tends to lose their bioactivity. In this study, we describe a procedure to immobilize human single-chain variable fragment (scFv) via genetic fusion to partial spider silk, which have a high tendency to adhere to solid supports. Two scFvs, directed towards serum proteins, were genetically fused to partial spider silk proteins and expressed as silk fusion proteins in E. coli. Antigen binding ability of scFvs attached to a partial silk protein denoted RC was investigated using microarray analysis, whereas scFvs fused to the NC silk variant was examined using nanoarrays. Results from micro- and nanoarrays confirmed the functionality of scFvs attached to both RC and NC silk, and also for binding of targets in crude serum. Furthermore, the same amount of added scFv gives higher signal intensity when immobilized via partial spider silk compared to when immobilized alone. Together, the results suggest that usage of scFv-silk fusion proteins in immunoassays could improve target detection, in the long run enabling novel biomarkers to be detected in crude serum proteomes. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/8152161
- author
- Thatikonda, Naresh ; Delfani, Payam LU ; Jansson, Ronnie ; Petersson, Linn LU ; Lindberg, Diana ; Wingren, Christer LU and Hedhammar, My
- organization
- publishing date
- 2015-10-16
- type
- Contribution to journal
- publication status
- published
- subject
- in
- Biotechnology Journal
- volume
- 11
- issue
- 3
- pages
- 437 - 448
- publisher
- John Wiley & Sons Inc.
- external identifiers
-
- pmid:26470853
- scopus:84959232625
- wos:000372141400016
- pmid:26470853
- ISSN
- 1860-6768
- DOI
- 10.1002/biot.201500297
- language
- English
- LU publication?
- yes
- id
- a51e8dd6-262e-41fe-a003-f6f01c66a938 (old id 8152161)
- date added to LUP
- 2016-04-01 10:27:09
- date last changed
- 2022-02-17 18:13:16
@article{a51e8dd6-262e-41fe-a003-f6f01c66a938,
abstract = {{Immobilizing biomolecules with retained functionality and stability on solid supports is crucial for generation of sensitive immunoassays. However, upon use of conventional immobilization strategies, a major portion of the biomolecules (e.g. antibodies) frequently tends to lose their bioactivity. In this study, we describe a procedure to immobilize human single-chain variable fragment (scFv) via genetic fusion to partial spider silk, which have a high tendency to adhere to solid supports. Two scFvs, directed towards serum proteins, were genetically fused to partial spider silk proteins and expressed as silk fusion proteins in E. coli. Antigen binding ability of scFvs attached to a partial silk protein denoted RC was investigated using microarray analysis, whereas scFvs fused to the NC silk variant was examined using nanoarrays. Results from micro- and nanoarrays confirmed the functionality of scFvs attached to both RC and NC silk, and also for binding of targets in crude serum. Furthermore, the same amount of added scFv gives higher signal intensity when immobilized via partial spider silk compared to when immobilized alone. Together, the results suggest that usage of scFv-silk fusion proteins in immunoassays could improve target detection, in the long run enabling novel biomarkers to be detected in crude serum proteomes.}},
author = {{Thatikonda, Naresh and Delfani, Payam and Jansson, Ronnie and Petersson, Linn and Lindberg, Diana and Wingren, Christer and Hedhammar, My}},
issn = {{1860-6768}},
language = {{eng}},
month = {{10}},
number = {{3}},
pages = {{437--448}},
publisher = {{John Wiley & Sons Inc.}},
series = {{Biotechnology Journal}},
title = {{Genetic fusion of single-chain variable fragments to partial spider silk improves target detection in micro- and nanoarrays.}},
url = {{http://dx.doi.org/10.1002/biot.201500297}},
doi = {{10.1002/biot.201500297}},
volume = {{11}},
year = {{2015}},
}