We report here a novel detection method in which the intrinsic affinity of proteins toward silver is used to induce Surface Enhanced Resonance Raman scattering (SERRS) of Raman active R6G dye conjugated to detection antibodies. We found a simple method of passive adsorption of silver nanoparticles onto PVDF membrane produces readily useful substrate for inducing the enhancement of Raman signal which could then be used to discern the presence of specific target molecules in quantitative manner. The effect is specific for PVDF membrane, other synthetic membranes such as nitrocellulose fail to produce the same effect. The method can be used for an accurate quantitative measurement of proteins labelled with Raman-active dyes. For example, it can be useful for quantification of detection antibodies eluted off after immunodetection of a target protein and subsequently allowed to adsorb onto silver substrate which can induce enhancement of Raman signal. Using mouse IgG molecules as a model antigen, adsorbed mIgG molecules were detected with limit of detection (3x S/N ratio over blank) down to 0.4ng (3 femtomoles) per well with greater than 1 log linear dynamic range. We also tested this method under sandwich immunodetection targeting c-peptide and show detection limit of 20 picogram (7 femtomoles).
- Copyright © 2011 by The American Association of Immunologists, Inc.