A new quantitative optical biosensor for protein characterisation.
A new optical biosensor is described based on a dual waveguide interferometric technique. By addressing the waveguide structure with alternate polarisations the optogeometrical properties (density and thickness) of adsorbed protein layers at the sensor (solid)-liquid interface have been determined. Differences in the waveguide mode dispersion between the transverse electric (TE) and transverse magnetic (TM) modes allow unique solutions for adlayer thickness and refractive index to be determined at all stages during the formation process. The technique has been verified using standard protein systems and by comparing the data with published work using X-ray crystallography and neutron reflection techniques. The data obtained was found to be in excellent agreement with previously reported X-ray experiments given that typical film thicknesses for streptavidin layers were in the range 5.5-6.5 nm compared with the short axis crystal structure of between 4.8 and 5.6 nm. The precision of the measurements taken was of the order of 40 pm with respect to adsorbed adlayer thicknesses. This biosensor approach provides measurements of both thickness and density of adlayers to a high precision, simultaneously and in real time enabling detail of the structure and function of proteins to be elucidated. From such data it is possible to obtain information on the orientation, distortion and efficiency of immobilisation procedures as well as the interaction event of interest. The technique is expected to find utility with those interested in protein structure and function. This is an area of growing importance within the life sciences as the demand for quantitative analytical techniques increases with the growth in "proteomics".
Cross
Graham H
GH
Department of Physics, University of Durham, South Road, Durham DH1 3LE, UK.
Reeves
Andrew A
AA
Brand
Stuart
S
Popplewell
Jonathan F
JF
Peel
Louise L
LL
Swann
Marcus J
MJ
Freeman
Neville J
NJ
D000906:Antibodies / Q000032:analysis / Q000737:chemistry; D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods; D001710:Biotin / Q000032:analysis / Q000737:chemistry; D004867:Equipment Design; D019544:Equipment Failure Analysis; D006801:Humans; D007368:Interferometry / Q000295:instrumentation* / Q000379:methods; D055095:Optics and Photonics / Q000295:instrumentation*; D011506:Proteins / Q000032:analysis* / Q000737:chemistry*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D019809:Streptavidin / Q000032:analysis / Q000737:chemistry
D000906:Antibodies; D011506:Proteins; D001710:Biotin; D019809:Streptavidin