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AbstractEffective detection of bio‐molecules relies on the precise design and preparation of materials, particularly in laser desorption/ionization mass spectrometry (LDI‐MS). Despite significant advancements in substrate materials, the performance of single‐structured substrates remains suboptimal for LDI‐MS analysis of complex systems. Herein, designer Au@SiO2@ZrO2 core‐shell substrates are developed for LDI‐MS‐based early diagnosis and prognosis of pancreatic cancer (PC). Through controlling Au core size and ZrO2 shell crystallization, signal amplification of metabolites up to 3 orders is not only achieved, but also the synergistic mechanism of the LDI process is revealed. The optimized Au@SiO2@ZrO2 enables a direct record of serum metabolic fingerprints (SMFs) by LDI‐MS. Subsequently, SMFs are employed to distinguish early PC (stage I/II) from controls, with an accuracy of 92%. Moreover, a prognostic prediction scoring system is established with enhanced efficacy in predicting PC survival compared to CA19‐9 (p < 0.05). This work contributes to material‐based cancer diagnosis and prognosis.