We present a strategy for the synthesis of multiplexed spectral encoder beads based on combinations of different surface enhanced Raman (SERS) signatures generated by dye-functionalized Ag nanoparticle tags. A key problem in SERS-based multiplexing arises in balancing the competitive binding of different signal generating dyes to the nanoparticle surfaces, which leads to difficulty in generating final summation spectra by design. We avoid this complication by decoupling the formation of individual tags from multiplexing of their spectra by self-assembly of different tag combinations onto SiO_(2) microbead supports via biotin-avidin binding. Linear combinations of individual nanoparticle tag spectra are generated in precursor solutions and are found to directly translate to the final encoder bead fingerprint spectrum in a 1:1 binding stoichiometry that preserves the original solution ratios. The result is an ability to multiplex spectral signatures in both frequency and intensity space to generate a large number of unique encoder signatures from a limited number of initial tag spectra. Raman microscopy of 75 individual beads shows that spectral response is highly uniform from bead-to-bead, making the encoder assemblies suitable for highly multiplexed bioassay applications and as model systems for cellular surface labeling studies for imaging and immunoassays.
展开▼
