Abstract: Objective: To establish a self-assembled nucleic acid aptamer probe technique for the visual detection of peripheral blood (1,3)-β-D-glucans in deep fungal infection. Methods: The (1,3)-β-D-glucans from Candida albicans were extracted by the alkali enzymatic method and digested by glucanase in vitro. The obtained water-soluble (1,3)-β-D-glucans were used as the target, and the single-strained DNA (ssDNA) aptamers which could specifically recognize and bind with glucans were screened from the single-strained DNA library using the systematic evolution of ligands by exponential enrichment (SELEX). The selected aptamers were designed to form the G-quadruplex structure containing hemin, which had the characteristics of peroxidase, and could catalyze the color development of TMB substrate for the color change judged by naked eyes. The (1,3)-β-D-glucan levels in 149 serum samples were detected by the established method and G-test, respectively, and their detection efficiency was evaluated. Results: The (1,3)-β-D-glucans were extracted from Candida albicans successfully, and the water-soluble (1,3)-β-D-glucans were obtained after digesting with glucanase. After SDS-PAGE separation and glycogen staining, it was found that water-soluble (1,3)-β-D-glucans were mainly distributed in three regions with relative molecular weight less than 1 700, about 4 600 and 10 000-15 000. The proportion of (1,3)-β-D-glucans with relative molecular weight less than 10 000 was higher, so they were used as the target for screening aptamers. After 8 rounds of positive screening and 4 rounds of negative screening, the last round of ssDNA library was amplified into dsDNA and cloned into pUC19 plasmid. The relative binding capacity of the aptamers amplified from the single positive clone was compared, and 6 aptamers with high binding capacity, named A1-A6 respectively, were obtained. The results of competitive experiments showed that six aptamers could recognize four different sites. The affinity between aptamers with high binding capacity and different concentrations of (1,3)-β-D-glucans was determined, and the results showed that the detection limit with naked eyes was 3.125 pg/mL, and that the linear range was between 1.6 pg/mL and 400 pg/mL. The (1,3)-β-D-glucans detection results of 149 serum samples showed that the detection performance of the aptamer self-assembled color system was better than that of G test (χ²=4.373, P=0.036 5). Conclusion: The visual nucleic acid aptamer probe is constructed successfully to detect serum (1,3)-β-D-glucans, which is expected to develop a point-of-care testing kit for the detection of deep fungal infection.