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Quantification of interfacial interactions between particles provides a way to regulate the interface behaviors of particles related with adhesion, aggregation, flotation, flocculation, membrane fouling, etc. Existing methods are based on assumptions of smooth particles although real particle surfaces are rather rough. This study proposed a new method to quantify interfacial interactions between two rough particles. In this study, a rigorous mathematical equation was firstly introduced to construct surface topography. In the framework of surface element integration (SEI) method, the spatial relationship between two rough particles was significantly explored, resulting in establishment of a formula of double integrals for interaction quantification. Thereafter, surface properties of the microbial aggregations obtained from a membrane bioreactor (MBR) were experimentally measured. With these data, the interfacial interactions between two rough microbial aggregations were numerically quantified according to composite Simpson’s rule. The new method was compared with Derjaguin approximation (DA) method. It was found that ripple frequency and particle radius had profound effects on the total interfacial interaction. This method has extensive application foreground in interfacial behavior research. [ABSTRACT FROM AUTHOR]