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A methodology based on species-unspecific isotope dilution-gel electrophoresis-laser ablation coupled with inductively coupled plasma mass spectrometry (GE-LA-ID-ICP-MS) was established the first time for the absolute quantification of proteins in human serum.The created species-unspecific isotope dilution in GE-LA-ICP-MS had achieved sufficient isotope exchange and isotopic equilibration successfully by mixing sulfur spike with protein strips.According to isotope dilution equation, the method was applied to a fast, accurate absolute quantification of transferrin and albumin in human serum.