Study on the mechanism of Bioelectric Recognition Assay: evidence for immobilized cell membrane interactions with viral fragments.
The Bioelectric Recognition Assay (BERA) is a whole-cell based biosensing system that detects the electric response of cultured cells, suspended in a gel matrix, to various ligands, which bind to the cell and/or affect its physiology. Previous studies have demonstrated the potential application of this method for rapid, inexpensive detection of viruses in a crude sample. However, the understanding, so far, of the fundamental processes that take place during cell-virus interactions within the probe has been rather limited. In the present study, we combined electrophysiological and fluorescence microscopical assays, so that we can prove that animal and plant cells immobilized in BERA sensors respond to different viruses primarily by changing their membrane potential. The response of immobilized cells against different viruses did not depend on the virus ability to penetrate the cell, but was modified after binding each virus to a virus-specific antibody or removal of its coat protein after treatment with a protease. Consequently, we were able to assay the presence of a virus in its complete form or fragments thereof. Combination of immunological recognition with the electrophysiological response of immobilized cells allows for a considerable increase of the specificity of the BERA biosensory assay. In addition, rather than simply detect the presence of a protein or genomic sequence, the method can help gain information on the bioactivity of a virus.
Kintzios
S
S
Laboratory of Plant Physiology, Agricultural University of Athens, Iera Odos 75, 11855, Athens, Greece. skin@aua.gr
Bem
F
F
Mangana
O
O
Nomikou
K
K
Markoulatos
P
P
Alexandropoulos
N
N
Fasseas
C
C
Arakelyan
V
V
Petrou
A-L
AL
Soukouli
K
K
Moschopoulou
G
G
Yialouris
C
C
Simonian
A
A
D000818:Animals; D001681:Biological Assay / Q000379:methods*; D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods; D018914:Cells, Immobilized / Q000502:physiology / Q000821:virology; D002522:Chlorocebus aethiops; D004563:Electrochemistry / Q000295:instrumentation / Q000379:methods*; D018259:Herpesvirus 1, Human / Q000302:isolation & purification* / Q000502:physiology; D007118:Immunoassay / Q000295:instrumentation / Q000379:methods*; D008564:Membrane Potentials / Q000502:physiology*; D010942:Plant Viruses / Q000302:isolation & purification* / Q000502:physiology; D011523:Protoplasts / Q000821:virology*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014709:Vero Cells