Tags

Type your tag names separated by a space and hit enter

Properties of a panel of single chain variable fragments against Potato leafroll virus obtained from two phage display libraries.
J Virol Methods. 1999 Aug; 81(1-2):159-68.JV

Abstract

Twelve single chain variable fragment (scFv) antibodies that bind to particles of Potato leafroll virus (PLRV) were obtained from two naive phage display libraries. Phages were selected against PLRV particles or dissociated PLRV particles immobilised onto tubes. Individual PLRV-binding scFv were identified by ELISA, after their expression either fused to the surface of phage particles, or as soluble scFv (scFv-c-myc), or as scFv-alkaline phosphatase fusion proteins (scFv-AP), obtained by subcloning into pSKAP/S. These procedures resulted in the isolation of scFv with different properties. For example, some of the scFv reacted strongly with virus particles but not with dissociated capsid protein, which suggests that they had reacted with discontinuous epitopes. Others reacted with dissociated capsid proteins and SDS-denatured protein, which suggests that they had reacted with continuous epitopes. ScFv were also subcloned into pC(L) for expression as fusion proteins with human kappa constant region (scFv-C(L)). Expression of these constructs in Escherichia coli yielded 0.2-1 mg protein per litre of bacterial culture. The different scFv fusion proteins were evaluated in ELISA to detect PLRV in leaf extracts of Physalis floridana. Absorbance values obtained with the fusion proteins were greater than those obtained with the scFv-c-myc, and were similar to those obtained in assays done using monoclonal or polyclonal antibodies.

Authors+Show Affiliations

Scottish Crop Research Institute, Dundee, UK.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

10488774

Citation

Harper, K, et al. "Properties of a Panel of Single Chain Variable Fragments Against Potato Leafroll Virus Obtained From Two Phage Display Libraries." Journal of Virological Methods, vol. 81, no. 1-2, 1999, pp. 159-68.
Harper K, Toth RL, Mayo MA, et al. Properties of a panel of single chain variable fragments against Potato leafroll virus obtained from two phage display libraries. J Virol Methods. 1999;81(1-2):159-68.
Harper, K., Toth, R. L., Mayo, M. A., & Torrance, L. (1999). Properties of a panel of single chain variable fragments against Potato leafroll virus obtained from two phage display libraries. Journal of Virological Methods, 81(1-2), 159-68.
Harper K, et al. Properties of a Panel of Single Chain Variable Fragments Against Potato Leafroll Virus Obtained From Two Phage Display Libraries. J Virol Methods. 1999;81(1-2):159-68. PubMed PMID: 10488774.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Properties of a panel of single chain variable fragments against Potato leafroll virus obtained from two phage display libraries. AU - Harper,K, AU - Toth,R L, AU - Mayo,M A, AU - Torrance,L, PY - 1999/9/17/pubmed PY - 1999/9/17/medline PY - 1999/9/17/entrez SP - 159 EP - 68 JF - Journal of virological methods JO - J. Virol. Methods VL - 81 IS - 1-2 N2 - Twelve single chain variable fragment (scFv) antibodies that bind to particles of Potato leafroll virus (PLRV) were obtained from two naive phage display libraries. Phages were selected against PLRV particles or dissociated PLRV particles immobilised onto tubes. Individual PLRV-binding scFv were identified by ELISA, after their expression either fused to the surface of phage particles, or as soluble scFv (scFv-c-myc), or as scFv-alkaline phosphatase fusion proteins (scFv-AP), obtained by subcloning into pSKAP/S. These procedures resulted in the isolation of scFv with different properties. For example, some of the scFv reacted strongly with virus particles but not with dissociated capsid protein, which suggests that they had reacted with discontinuous epitopes. Others reacted with dissociated capsid proteins and SDS-denatured protein, which suggests that they had reacted with continuous epitopes. ScFv were also subcloned into pC(L) for expression as fusion proteins with human kappa constant region (scFv-C(L)). Expression of these constructs in Escherichia coli yielded 0.2-1 mg protein per litre of bacterial culture. The different scFv fusion proteins were evaluated in ELISA to detect PLRV in leaf extracts of Physalis floridana. Absorbance values obtained with the fusion proteins were greater than those obtained with the scFv-c-myc, and were similar to those obtained in assays done using monoclonal or polyclonal antibodies. SN - 0166-0934 UR - https://www.unboundmedicine.com/medline/citation/10488774/Properties_of_a_panel_of_single_chain_variable_fragments_against_Potato_leafroll_virus_obtained_from_two_phage_display_libraries_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0166-0934(99)00071-3 DB - PRIME DP - Unbound Medicine ER -