Abstract Hierarchical mesoporous silica (meso-SiO2) films with 90nm thickness and 4.34nm interpore spacing were fabricated on the SPR sensor chips for rapid detection of small and large molecules. With non-adsorbing glucose molecules the refractive-index sensitivity of the sensor was detected to increase from 8283.2nm/RIU to 22354nm/RIU with moving the resonance wavelength from λR=920nm to λR=1220nm. Adsorption of CTAB molecules within the meso-SiO2 film leads to a more than 7 times enhancement in the sensor response, showing the Langmuir isotherm adsorption behavior. The sensor can also detect proteins larger than the pore size of the meso-SiO2 film, whose rough surface makes the sensor responses very close to those obtained with the uncoated SPR chips. The findings indicate that the nonspecific adsorption of goat anti-human IgG molecules on the meso-SiO2 film did not bring about obvious sterical hindrance to their proper specificity, enabling the sensor to detect human IgG at a concentration as low as 10nM. The work demonstrated the outstanding applicability of the meso-SiO2 film coated SPR sensors for real-time and sensitive detection of both small and large molecules.