title: Recombinant single-chain variable fragment and single domain antibody piezoimmunosensors for detection of HIV1 virion infectivity factor.
abstract: In this paper recombinant single-chain fragments (scFv-4BL), and single domain antibodies (4BL-V(H)) and (4BL-V(H)D) generated against HIV1 virion infectivity factor (Vif) are used to develop piezoimmunosensors for HIV1 recognition. Mixed self assembled monolayers were generated at the surface of gold coated crystal sensors to which scFv-4BL, 4BL-V(H), or 4BL-V(H)D were immobilized. Impedance analysis was used to discriminate interfering signals from frequency variation data and to increase the sensor sensitivity. The elimination of interfering signals enabled the quantification of the amount of immobilized protein and gave some indication on the viscoelasticity of immobilized biofilms. All the modified sensors were able to specifically recognize HIV1 Vif in liquid samples. The results indicate that lower sensitivities are obtained with 4BL-V(H) single domain antibodies, possibly due to its higher hydrophobic character. The sensitivity obtained when using scFv-4BL was reestablished when using the more hydrophilic 4BL-V(H)D single domain. 4BL-V(H)D piezoimunosensors were effective in recognizing HIV1 Vif from protein mixtures and from cell extracts of human embryonic kidney cells expressing HIV1 Vif. The results presented in this paper demonstrate the potential applicability of the developed piezoimmunosensors to monitor HIV1 infection evolution.
authors.affiliation: IBB-Institute for Biotechnology and Bioengineering, Centro de Biomedicina Molecular e Estrutural, Universidade do Algarve, 8000 Faro, Portugal. gferrei@ualg.pt
authors.forename: Guilherme N M
authors.initials: GN
authors.lastname: Ferreira
authors.forename: João M
authors.initials: JM
authors.lastname: Encarnação
authors.forename: Luis
authors.initials: L
authors.lastname: Rosa
authors.forename: Rogério
authors.initials: R
authors.lastname: Rodrigues
authors.forename: Roberta
authors.initials: R
authors.lastname: Breyner
authors.forename: Sara
authors.initials: S
authors.lastname: Barrento
authors.forename: Luisa
authors.initials: L
authors.lastname: Pedro
authors.forename: Frederico
authors.initials: F
authors.lastname: Aires da Silva
authors.forename: João
authors.initials: J
authors.lastname: Gonçalves
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D002460:Cell Line; D015483:HIV Antibodies / Q000276:immunology*; D006801:Humans; D007118:Immunoassay / Q000379:methods*; D007135:Immunoglobulin Variable Region / Q000276:immunology*; D011994:Recombinant Proteins / Q000276:immunology / Q000302:isolation & purification; D054320:vif Gene Products, Human Immunodeficiency Virus / Q000032:analysis* / Q000276:immunology / Q000302:isolation & purification
chemical_list: D015483:HIV Antibodies; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins; D054320:vif Gene Products, Human Immunodeficiency Virus; C514847:vif protein, Human immunodeficiency virus 1