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Methodologies play a crucial role in driving scientific advancement. Mass spectrometric techniques have been integrated into our skill set for protein structure analysis. My laboratory has pioneered novel analytical methods based on the elution properties of peptide products from protein digestion, uncovering cellular protein modifications such as phosphorylation and acetylation for many groups worldwide. My subsequent experience in the biotechnology industry further inspired me to develop innovative methods that enable the simultaneous screening of multiple types of protein modifications in a single analysis. Despite these progresses in protein modification analysis, we have not yet achieved our original goal of identifying plasma protein modifications as disease biomarkers. Around a decade ago, this challenge led us to explore the potential of protein complexes as long-sought disease biomarkers. With our in-depth knowledge about plasma protein structures, we developed platform technologies to profile proteins present in liquid chromatography fractions from the tested plasma sample. These efforts have proven to be a robust approach, quickly yielding several complex biomarkers for various cancers. Furthermore, these protein structural alterations can be quantitatively assessed using automated western blotting analyses, shaping the future direction for developing clinical diagnostic tests aimed at high-sensitivity screening for diseases, such as early-stage cancers.