Comparison of surface-enhanced resonance Raman scattering and fluorescence for detection of a labeled antibody.
A comparison is made of the quantitative detection of a labeled antibody by surface-enhanced resonance Raman scattering (SERRS) and by fluorescence using the same instrument with the same laser excitation source. The area under the curve for the fluorescence band is greater than for any single peak in the SERRS spectrum, but the broad fluorescence band is more difficult to discriminate from the background at low concentrations. Using the peak height of one SERRS band and the peak height at the fluorescence maximum, the detection limit for SERRS was lower (1.19 x 10-11 mol.dm-3) than that obtained using fluorescence (3.46 x 10-10 mol.dm-3). The SERRS detection limit is calculated for the concentration of the sample added, but compared to fluorescence, there is an additional dilution step due to the addition of the colloid and the extent of this dilution is dependent on assay format. For comparison with the detection limits found earlier with labeled oligonucleotides, SERRS was remeasured with a 10 s accumulation time, and the final concentration in the cuvette after colloid addition and before any adsorption to the silver was used to calculate a detection limit of 2.79 x 10-13 mol.dm-3. This is comparable to the detection limit found using a similar SERRS procedure for an oligonucleotide labeled with the same dye. This experiment is dependent on many parameters that could affect this result, including the nature of the SERRS substrate, the excitation wavelength, and the dye chosen. However, the result indicates that SERRS can give assay sensitivities comparable or better than fluorescence for quantitative direct assay determination, suggesting that the much greater potential for multiple analyte detection could be exploited.
Sabatté
Gwénola
G
Department of Pure & Applied Chemistry, University of Strathclyde, 298 Cathedral Street, Glasgow, G1 1XL, UK.
Keir
Ruth
R
Lawlor
Margaret
M
Black
Murdo
M
Graham
Duncan
D
Smith
W Ewen
WE
D000906:Antibodies / Q000032:analysis* / Q000737:chemistry / Q000276:immunology*; D015394:Molecular Structure; D013050:Spectrometry, Fluorescence / Q000379:methods*; D013059:Spectrum Analysis, Raman / Q000379:methods*; D013499:Surface Properties
D000906:Antibodies