Detection of pathogenic bacteria using a homogeneous immunoassay based on shear alignment of virus particles and linear dichroism.
Biomolecular detection has for a long time depended on a relatively small number of established methodologies. Many of these depend on the detection of a ligand-antibody complex using some kind of optical technique, e.g., chemiluminescence. Before this measurement can be made, the ligand-antibody complex generally has to be separated from bulk contaminants. This process involves the implementation of a heterogeneous assay format involving immobilization of the antibody onto a solid support to enable washing of the unbound ligand. This approach has a number of inherent issues including being slow and complex and requiring the use of expensive reagents. In this paper, we demonstrate how we have harnessed a biologically inspired nanoparticle to provide the basis for a homogeneous assay which requires no immobilization. The method relies on using fluid shear flow to align a fiber-like nanoparticle formed from a filamentous virus, M13, combined with a ligand-specific antibody. This renders the particle visible to linear dichroic spectroscopy, which provides an easily interpretable signal. The addition of the target ligand (in this case Escherichia coli O157) leads to the formation of a nanoparticle-ligand particle that is unable to align, leading to the perturbation of the linear dichroism signal.
School of Biosciences, University of Birmingham, Edgbaston, Birmingham, West Midlands, B15 2TT, UK.
Raúl
R
Pacheco-Gómez
Julia
J
Kraemer
Susan
S
Stokoe
Hannah J
HJ
England
Charles W
CW
Penn
Emma
E
Stanley
Alison
A
Rodger
John
J
Ward
Matthew R
MR
Hicks
Timothy R
TR
Dafforn
D000906:Antibodies / Q000737:chemistry; D001419:Bacteria / Q000302:isolation & purification* / Q000472:pathogenicity; D007118:Immunoassay / Q000379:methods*; D008024:Ligands; D049449:Luminescence; D013057:Spectrum Analysis / Q000379:methods*; D014771:Virion
D000906:Antibodies; D008024:Ligands