The effects of concentration and type of anions on aggregation of Congo red in solution were studied with UV-Visible spectroscopy. And the influence of anions on the complex of Congo red and oat β-glucan was further studied. The results showed that the maximum absorption wavelength of Congo red in aqueous solution shifted toward the shorter wavelength, and the absorbency lowered considerably with increasing ionic concentration. There was a significant linear correlation between logarithm of anion concentration and the maximum absorption wavelength, with the peak absorbance, and the absorbance at 499 nm. The influence of anions on aggregation of Congo red was in accordance with Hofmeister series, indicating that hydrophobic interaction is an important driving force for Congo red molecules to assemble into micelles. As for the complexation of Congo red and oat β-glucan, when the concentration of anions exceeded the first critical concentration, the micelle of Congo red began to form and then combined to β-glucan. The absorption peak of the complex presented at 556 nm. When the concentration of anions exceeded the second critical concentration, the complex was developed into supramolecular structure through aggregation of Congo red micelles on Congo red/β-glucan complex, which resulted in the shift of absorption peak toward 583 nm. And the presence of Mie scattering effects at the longer wavelength indicated the formation of larger supramolecular structure. The effect of anions on the above supramolecular structures also followed the order of Hofmeister series, showing that the aggregation of Congo red/β-glucan complex into supramolecular structure was mainly driven through hydrophobic interactions between Congo red micelles on the complex. The research indicated that ions had an important effect on the aggregation of Congo red and the interaction between biopolymers and Congo red.