Recombinant single-chain variable fragment and single domain antibody piezoimmunosensors for detection of HIV1 virion infectivity factor.
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.
Ferreira
Guilherme N M
GN
IBB-Institute for Biotechnology and Bioengineering, Centro de Biomedicina Molecular e Estrutural, Universidade do Algarve, 8000 Faro, Portugal. gferrei@ualg.pt
Encarnação
João M
JM
Rosa
Luis
L
Rodrigues
Rogério
R
Breyner
Roberta
R
Barrento
Sara
S
Pedro
Luisa
L
Aires da Silva
Frederico
F
Gonçalves
João
J
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
D015483:HIV Antibodies; D007135:Immunoglobulin Variable Region; D011994:Recombinant Proteins; D054320:vif Gene Products, Human Immunodeficiency Virus; C514847:vif protein, Human immunodeficiency virus 1