Fig. 3

Quantitative aptaprobe-based sandwich assay (ALISA) for detecting GPC3. A Schematic overview of the GPC3 ALISA platform. The graph is shown for the ALISA combination in the panel as detection GPC3. C1, microwell plate; C2, non-specific binging of reporter aptaprobes; C3, non-specific target binding without capture aptaprobe; and two negative controls—C4, biotinylated GPC3_9 and fluorescein amidite (FAM)-labeled GPC3_3; and C5, biotinylated GPC3_2 and FAM-labeled GPC3_3. The sandwich ALISA [Test4] quantify GPC3 between two layers of aptaprobes (capture and detection aptaprobes, GPC3_2 and GPC3_3, respectively). [Test3] did not show the fluorescence signal as the immobilized GPC3_9 interrupt the signaled GPC3_3, due to the same binding site. B Linear regression of the GPC3 ALISA. All experiments were performed in triplicate. C 3D structure two aptaprobes (biotinylated GPC3_2 and FAM-labeled GPC3_3) bind to GPC3 simultaneously and without interference from each other, indicating recognition of distinct aptatope regions. All experiments were used the 1 μM/well of aptaprobes