Fig. 4: Fluorescence-based immunoassay.

a Kinetic curves related to (1) Ab-analyte and (2) analyte-aptamer binding dynamics. Both processes reach their equilibrium in a short time by incubation and tilting mixing (80 min and 45 min, respectively). The time difference is ascribable to the significantly larger PfLDH mass. The data are well fitted by exponential curves (orange and blue continuous lines). b Fluorescence images acquired at different PfLDH concentrations in spiked human blood. c Calibration curve (fluorescence intensity vs PfLDH concentration in spiked human blood) of the immunoassay for PfLDH concentration in the range 1 fM to 100 nM (35 fg/mL to 3.5 μg/mL). The data are best fitted by the four-parameter Hill equation (red solid line). The gray region represents the 3σ noise level recorded in uncontaminated human blood (LOD = 18 fM (0.6 pg/mL)). d Specificity of the immunoassay against PvLDH (90% residue identity with PfLDH) at a concentration of 100 nM in spiked human blood. No cross-reaction detected with the main PfLDH competitor highlights the extremely high specificity of the aptamers used as top bioreceptor layer (**p-value < 0.001). All the data underlying averaged value are presented as mean value ± SD and are representative of ten technical repeats. Source data are provided as a Source data file.