The efficient isolation and specific discrimination of circulating tumor cells (CTCs) is expected to provide valuable information for understanding tumor metastasis and play an important role in the treatment of cancer patients. In this study, we developed a novel and rapid method for efficient capture and specific identification of cancer cells using hyaluronic acid (HA)-modified SiO2-coated magnetic beads in a microfluidic chip. First, polyacrylamide-surfaced SiO2-coated magnetic beads (SiO2@MBs) were covalently conjugated with HA, and the created HA-modified SiO2@MBs (HA-SiO2@MBs) display binding specificity to HeLa cells (a human cervical carcinoma cell line) overexpressing CD44 receptors. After incubating the HA-SiO2@MBs with cancer cells for 1 h, the mixture of MBs and cells was drawn into a designed microfluidic channel with two inlets and outlets. Through the formation of lamellar flow, cells specifically bound with the HA-SiO2@MBs can be separated under an external magnetic field with a capture efficiency of up to 92.0%. The developed method is simple, fast, and promising for CTC separation and cancer diagnostics applications.