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Its application to clinical study enables simultaneous imaging\u2010based morphological observations and mass spectrometry analysis. Application of fully informative material like tissue allows obtaining the complex and unique profile of analyzed samples. This knowledge leads to diagnosing disease, studying the mechanism of cancer development, selecting the potential biomarkers as well as correlating obtained images with prognosis. Nevertheless, it is worth noticing that this method is found to be objective but the result of the analysis is mainly influenced by the sample preparation protocol, including the collection of biological material, its preservation, and processing. However, the application of this approach requires a special sample preparation procedure. The main goal of the study is to present the current knowledge on the clinical application of matrix\u2010assisted laser desorption\/ionization with imaging mass spectrometry in cancer research, with particular emphasis on the sample preparation step. 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