An effective technique for the suppression of matrix-related ions in matrix-assisted laser desorption/ionization (MALDI) mass spectrometry have been developed. Using typical organic matrices such as THAP (2,4,6-trihydroxyacetophenone) and CHCA (α-cyano-4-hydroxycinnamic acid) in a cyclodextrin cavity, we successfully measured the mass peaks of only protonated matrix ions and significantly suppressed their intensities and fragmentation. In addition, it became possible to analyze the mass peak of the analyte molecules (substance P and adenosine) without any interference from the matrix. We believe that this technique could be a powerful tool for MALDI mass spectrometry, particularly for low-molecular-weight compounds.