ORIGINAL ABSTRACT

Gold nanospheres (AuNPs) and gold nanoflowers (AuNFs) are widely used as platforms for DNA aptamer functionalization, while conjugation behavior and colloidal tolerance remain important factors affecting subsequent sensing-oriented optimization. In this study, 82-nt thiolated DNA aptamer constructs bearing either 3'-SH or 5'-SH terminal modification were immobilized onto citrate-stabilized AuNPs and AuNFs under matched stepwise salt-aging conditions. Apparent nanoparticle-associated DNA output was estimated by Qubit-based measurement of unbound ssDNA in the supernatant and expressed as mass-based loading output (ng). Under the tested stock-dispersion conditions, AuNP samples showed higher apparent conjugation output than AuNF samples. Specifically, the apparent conjugation yields for AuNPs were 80.65 ± 1.64% (3'-SH) and 84.76 ± 1.98% (5'-SH), whereas those for AuNFs were 66.64 ± 3.36% (3'-SH) and 73.65 ± 1.36% (5'-SH). The corresponding apparent DNA loading outputs were 2329.7 ± 47.4 ng and 2448.7 ± 57.1 ng for AuNPs, and 1925.1 ± 97.0 ng and 2127.4 ± 39.3 ng for AuNFs. DLS size increases and zeta potential shifts toward more negative values were consistent with the formation of a DNA-associated interfacial layer, while TEM images supported morphology retention after conjugation. A qualitative visual salt-challenge assessment indicated that aptamer-functionalized nanoparticles displayed improved resistance to salt-induced aggregation relative to bare particles under the tested conditions. Because the commercially supplied AuNP and AuNF dispersions were not normalized to identical particle number or accessible surface area, the reported values should be interpreted as comparative apparent outputs rather than intrinsic loading capacities. Within this scope, the present study provides a convenient preliminary materials-level evaluation of thiolated aptamer conjugation behavior and may support future glyphosate aptasensor optimization.