Features of the As389/HHQ-BSA ELISA for the detection of HQNO

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... these studies, the conditions established for the As389/HHQ-BSA ELISA improved significantly the detectability of the assay and involved the use of PBS-6.5 as the assay buffer. The calibration curve recorded under these conditions is shown in Fig. 2. HQNO could be detected with a LOD of 0.27 6 0.09 nM, an IC 50 of 4.20 6 0.86 nM, and a dynamic range compressed between 0.72 6 0.18 to 26.71 6 0.96 nM (Table 1) (Table 1). Each calibration point was measured in triplicates on the same ELISA plate and the results showed the average and standard deviation of analysis made on three different days. ...

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... these studies, the conditions established for the As389/HHQ-BSA ELISA improved significantly the detectability of the assay and involved the use of PBS-6.5 as the assay buffer. The calibration curve recorded under these conditions is shown in Fig. 2. HQNO could be detected with a LOD of 0.27 6 0.09 nM, an IC 50 of 4.20 6 0.86 nM, and a dynamic range compressed between 0.72 6 0.18 to 26.71 6 0.96 nM (Table 1) (Table 1). Each calibration point was measured in triplicates on the same ELISA plate and the results showed the average and standard deviation of analysis made on three different days. ...

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... the potential nonspecific interferences in this complex biological matrix were assessed by building calibration curves in MH diluted several times with PBS-6.5 (Fig. S2). As shown in Fig. 2 and Table 1, the assay performed well in undiluted MH broth, although compared to the assay run in buffer, a slight increase of the maximum signal with a concomitant decrease of the assay detectability (IC 50 4.2 in buffer versus 14.6 in MH) could be observed. This effect diminished when diluting the MH media with the assay buffer reaching better features at 1/5 dilution factor (IC 50 2.7 nM, 13.5 nM considering the dilution of the culture media with the assay buffer). ...

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... bioconjugates were purified by dialysis against 0.5 mM PBS (5 Â 5 L) and Milli-Q water (1 Â 5 L), and freeze-dried at 280°C. A small fraction (20 mL) of the HQNO-BSA was kept for MALDI-TOF analysis, rendering a hapten density of 21 haptens per molecule of BSA (Table S1). ...