We present the results of matrix-assisted laser desorption/ionization (MALDI) imaging and direct molecular identification using tandem mass spectrometry (MS/MS) in colon cancer liver metastasis. Cancer tissue was removed from a Japanese patient and frozen immediately without any fixations. The sections were sliced to a thickness of 3 microm. The matrix for lipid ionization was 2,6-dihydroxy acetophenone. The matrix solution was applied with an airbrush into a thin uniform matrix layer on the tissue surface. After two-dimensional laser scanning, the images were reconstructed as a function of m/z from a few hundred obtained spectra. In the obtained images, the existence of molecules was represented by a pseudo-color corresponding to the signal intensity. In a feasibility study, we picked up a localized signal, m/z 725 in a cancerous area. The MS/MS result suggested that m/z 725 was sphingomyelin(16:0)+Na. Thus, we successfully show the feasibility of MALDI imaging as a tool for the analysis of pathological specimens.