Microwave-triggered surface plasmon coupled chemiluminescence.

Surface plasmon spectroscopy (SPS) is commonly implemented as an analytical tool to characterize thin kinetic processes and binding reactions at interfaces.1 Due to the limited sensitivity of SPS for the detection of dilute analytes of low molecular mass, surface plasmon coupled fluorescence (SPCF) or, alternatively, surface plasmon coupled emission (SPCE) has been proposed as an alternative means to improve the sensitivity of the technique.2,3 While SPCF was initially implemented to detect isotropic fluorescence emission,2 SPCE demonstrated that the emission from randomly oriented fluorophores close to metal surfaces was polarized and directional through the back of the thin metal film.3 As a result, SPCE has facilitated improved detection of chromophores bound to thin metal continuous films.3 In addition to directional and polarized fluorescence emission, our group has also reported surface plasmon coupled phosphorescence (SPCP)4 and surface plasmon coupled chemiluminescence (SPCC).5,6 As a result, surface plasmon coupled phenomena can be implemented to improve the detectability of biomolecular interactions at surfaces from weakly emitting species. In recent reports, we also introduced a new technology, microwave-triggered metal-enhanced chemiluminescence (MT-MEC),7 to increase the detectability of surface-bound HRP enzyme. With MT-MEC, short microwave pulses triggered increased photon flux from HRPcatalyzed chemiluminescent reactions.8,9 More recently, we implemented planar aluminum structures to focus microwave fields in a conventional microwave cavity and, subsequently, locally “trigger” chemiluminescent reactions. Combining the advantages of SPCC, MT-MEC, and microwavefocused chemiluminescence (MFC) technologies, we now introduce a new platform technology, microwave-triggered surface plasmon coupled chemiluminescence (MT-SPCC). With this new technique, we present a proof-in-principle method that combines the benefits of directional and polarized plasmon coupled emission with a microwave technology that induces locally “triggered” chemiluminescence emission.10 As a result, this technique may be implemented to further facilitate high sensitivity detection for chemiluminescent based surface assays, such as immunoassays.9,11,12 In this paper, the proof-of-principle of this new technique, microwavetriggered surface plasmon coupled chemiluminescence (MT-SPCC), is presented. MT-SPCC experiments were performed using green emission chemiluminescent reagents as previously described.7 Surface plasmon coupled chemiluminescence was collected using an optical scheme shown in Figure 1. Briefly, a small (approximately 1 in.) opening was created in the base of a conventional 2.45 GHz microwave cavity, and an optical configuration was designed to maximize the collection efficiency of the directional chemiluminescence emission. Pulsed SPCC intensities can be detected with a high sensitivity CCD (Retiga-SRV, QImaging, Inc.) or a fiber coupled to a spectrophotometer to collect time-dependent emission spectra with a time resolution of 100 ms (Figure 1, bottom right). Sample geometries were arranged as shown in Figure 1. In order to demonstrate the feasibility of localized MT-SPCC, 2.5 mm equilateral triangles, Ag continuous films 50 nm thick were vapor deposited on glass substrates with and without continuous silver films approximately 40 nm thick (Figure 2, top middle). The “bowtie” configuration was achieved by aligning the apex of an inverted 2.5 mm triangle with the tip of the mirror triangle, such that the tips are separated by a gap distance of 1 mm (Figure 2, top right). Imaging chambers were affixed to the center of 5 × 5 mm substrates and proximal to apexes of substrates modified with Ag triangles 50 nm thick. Chambers were subsequently filled with 20 μL of green chemiluminescence solution. The substrates were affixed to a glass prism using index matching immersion oil. Samples were positioned relative to the optical scheme at the base of the microwave to maximize the collection efficiency of the broad angular distribution (∼47-70°) inherent in plasmon coupled directional chemiluminescence.5,6 Figure 1. Microwave-triggered surface plasmon coupled chemiluminescence (MT-SPCC) optical scheme (left). Sample geometry for MT-SPCC experiments (top right) and typical MT-SPCC result (bottom right).