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This letter reports the measurement of single living cells’ refractive index (RI) using an on-chip fiber-based Fabry-Pérot cavity by a differential method. In experiment a single cell is captured into the cavity, then the spectral shift in response to the buffer change and the cell presence/absence can be used to determine the cell’s RI and size. Experiment on kidney cancer cells measures an effective RI of 1.399 at 0.1% accuracy. Compared with other approaches, the differential method eliminates uncertain factors and thus ensures high accuracy. The microchip facilitates automatic detection and makes it promising for label-free drug screening.