The paper presented a novel multiplex chemiluminescence (CL) immunoassay method, which was based on silica colloidal crystals beads (SCCBs) as carriers using enhanced chemiluminescence (CL) detection of a horseradish peroxidase (HRP) catalyzing the luminal reaction. The monodisperse and size-controlled SCCBs were fabricated by a microfluidic device. The experimental conditions were optimized and analytical performance was studied. Results showed that the SCCBs as supports were much sensitive (0.05 ng/ml IgG) than the glass beads (18 ng/ml IgG) and the planar carriers (125 ng/ml IgG). A multiplex immunoassay suggested the feasibility of high throughput applications. This novel immunoassay system provided a simple, sensitive and low-cost approach for multianalyte immunoassay without using of expensive array detector.