This study describes a new means to conduct molecular recognition-based analysis using mass spectrometry. Taking advantage of the unique characteristic of the absorption capacity of the TiO2 sol-gel material in the UV region, a TiO2 sol-gel-deposited thin film was employed as the sample substrate to assist in UV laser desorption/ionization of analytes. Sol-gels are polymeric materials that are easy to prepare and modify at low temperatures. Molecularly imprinted TiO2 sol-gels were generated for molecular recognition-based analysis. α-Cyclodextrin (CD) was selected as the template molecule and doped into TiO2 in a sol-gel reaction. The molecularly imprinted TiO2 sol was spin-coated on a glass slide, and appropriate template cavities in the TiO2 sol-gel material were formed after the template molecules were removed. We demonstrate that this modified glass slide can be used to select α-CD from a sample solution containing equal amounts of α-, β-, and γ-CD (50 ppb each, 18 mL); α-CD was directly detected from the modified glass slide by matrix-assisted laser desorption/ionization mass spectrometry without the addition of extra matrix. This approach provides a new detection method for molecular recognition-based analysis.