Of the immobilized antibodies and lowered mass transfer resistance. The immobilized antibodies were distributed uniformly within the particles, which facilitated the approach of immunoreagents and improved the kinetics of the antibody-antigen interaction. This improved experimental layout allowed us to obtain a high sensitivity SCCB suspension array. To apply the photonic suspension array, a simple platform was developed by incorporating a fiber optic spectrometer into a microscope for decoding and detection of the SCCBs (Fig. S4 in the Supporting information). When the SCCBs were exposed to white light under normal incidence through the microscope, the reflection peaks could be detected and the peak positions recorded for decoding. The fluorescence signals could be measured by replacing the input white light with blue light at a wavelength of 488 nm. Fig. 1A and Fig. 1B show images of multiplex detection obtained under 16574785 HIF-2��-IN-1 site bright and dark fields, respectively. Prior to multiple analysis, it was essential to investigate the cross-reactivity between the two immobilized antibodies and both FF competitors and FC competitors. When each type of modified SCCBs was incubated with a single competitor (FF competitors or FC competitors), the suspension array system showed only a singleAccuracy (Analysis of Spiked Samples)Grape, lettuce and cabbage from a local market and tap water from our laboratory were chosen for recovery studies. CLT and FNT standard stock solutions were prepared at 10, 100 and 1000 ng/mL. Grape peel, lettuce and cabbage leaves were chopped into fine pieces and 1 mL of each solution was added to 1 g of the samples. After standing overnight at 4uC for 24 h, the samples were shaken in 5 mL of methanol for 1 h and then filteredFigure 2. Response of the suspension array system to both competitors. Fluorescence intensity of the two types of SCCB modified by anti-FNT antibody and anti-CLT antibody, respectively, for FF competitors (a) or FC competitors (b) or a mixture of FF and FC competitors; the red and green bars represent the response of the system to FF and FC competitors, respectively. doi:10.1371/journal.pone.0066703.gDetection of Pesticides with a Suspension ArrayFigure 3. Optimization of experimental conditions and standard curves. Chebulagic acid effects of different amounts of mouse monoclonal antibodies (A) and competitors (B) on fluorescence 23727046 intensities, effects of incubation time on fluorescence intensities (C) and standard curves of the photonic suspension array (D). Each point was obtained by detecting 5 SCCBs. doi:10.1371/journal.pone.0066703.gresponse, corresponding to its competitor, with no response for the other competitor (Fig. 2a and b), indicating negligible crossreactivity. When incubated with both FF competitors and FC competitors, our suspension array showed responses to both, indicating that the two antibodies were successfully immobilized onto the surface of SCCBs (Fig. 2c). Hence, the suspension array system could be used to detect the selected pesticides, simultaneously.Optimization of Competitive Fluorescence ImmunoassayThe amounts of antibodies and competitors added and the incubation time affected the sensitivity of our suspension array. In Fig. 3A, differences are shown between the fluorescence intensities and the amount of antibodies added. With increasing addition of antibodies, the fluorescence intensities increased and showed clear points of inflection. Hence, optimal amounts of the two antibodies were chosen as.Of the immobilized antibodies and lowered mass transfer resistance. The immobilized antibodies were distributed uniformly within the particles, which facilitated the approach of immunoreagents and improved the kinetics of the antibody-antigen interaction. This improved experimental layout allowed us to obtain a high sensitivity SCCB suspension array. To apply the photonic suspension array, a simple platform was developed by incorporating a fiber optic spectrometer into a microscope for decoding and detection of the SCCBs (Fig. S4 in the Supporting information). When the SCCBs were exposed to white light under normal incidence through the microscope, the reflection peaks could be detected and the peak positions recorded for decoding. The fluorescence signals could be measured by replacing the input white light with blue light at a wavelength of 488 nm. Fig. 1A and Fig. 1B show images of multiplex detection obtained under 16574785 bright and dark fields, respectively. Prior to multiple analysis, it was essential to investigate the cross-reactivity between the two immobilized antibodies and both FF competitors and FC competitors. When each type of modified SCCBs was incubated with a single competitor (FF competitors or FC competitors), the suspension array system showed only a singleAccuracy (Analysis of Spiked Samples)Grape, lettuce and cabbage from a local market and tap water from our laboratory were chosen for recovery studies. CLT and FNT standard stock solutions were prepared at 10, 100 and 1000 ng/mL. Grape peel, lettuce and cabbage leaves were chopped into fine pieces and 1 mL of each solution was added to 1 g of the samples. After standing overnight at 4uC for 24 h, the samples were shaken in 5 mL of methanol for 1 h and then filteredFigure 2. Response of the suspension array system to both competitors. Fluorescence intensity of the two types of SCCB modified by anti-FNT antibody and anti-CLT antibody, respectively, for FF competitors (a) or FC competitors (b) or a mixture of FF and FC competitors; the red and green bars represent the response of the system to FF and FC competitors, respectively. doi:10.1371/journal.pone.0066703.gDetection of Pesticides with a Suspension ArrayFigure 3. Optimization of experimental conditions and standard curves. Effects of different amounts of mouse monoclonal antibodies (A) and competitors (B) on fluorescence 23727046 intensities, effects of incubation time on fluorescence intensities (C) and standard curves of the photonic suspension array (D). Each point was obtained by detecting 5 SCCBs. doi:10.1371/journal.pone.0066703.gresponse, corresponding to its competitor, with no response for the other competitor (Fig. 2a and b), indicating negligible crossreactivity. When incubated with both FF competitors and FC competitors, our suspension array showed responses to both, indicating that the two antibodies were successfully immobilized onto the surface of SCCBs (Fig. 2c). Hence, the suspension array system could be used to detect the selected pesticides, simultaneously.Optimization of Competitive Fluorescence ImmunoassayThe amounts of antibodies and competitors added and the incubation time affected the sensitivity of our suspension array. In Fig. 3A, differences are shown between the fluorescence intensities and the amount of antibodies added. With increasing addition of antibodies, the fluorescence intensities increased and showed clear points of inflection. Hence, optimal amounts of the two antibodies were chosen as.
