A protein of a biological sample is usually quantified by immunological techniques based on antibodies. Mass spectrometry offers alternative approaches that are not dependent on antibody affinity and avidity, protein isoforms, quaternary structures, or steric hindrance of antibody-antigen recognition in case of multi-protein complexes. One approach is the use of stable isotope-labeled internal standards, another the direct exploitation of mass spectrometric signals recorded by LC-MS/MS analysis of protein digests. Here, we assessed peptide match score summation index (PMSSI) based on probabilistic peptide scores calculated by the PHENYX protein identification engine for absolute protein quantification, in accordance with protein abundance index PAI as proposed by Mann and colleagues. Using synthetic protein mixtures, we demonstrated that this approach works well, although proteins can have different response factors. Applied to high-density lipoproteins (HDL), this new approach compared favorably to alternative protein quantitation methods like UV detection of protein peaks separated by capillary electrophoresis, or quantitation of protein spots on SDS-PAGE. We compared the protein composition of a well defined HDL density class isolated from plasma of seven hypercholesterolemia subjects having low or high HDL cholesterol with HDL from nine normolipidemia subjects. The quantitative protein patterns distinguished individuals according to the corresponding concentration and distribution of cholesterol from serum lipid measurements of the same samples, and revealed that hypercholesterolemia in unrelated individuals is the result of different deficiencies. The presented approach is complementary to HDL lipid analysis, does not rely on complicated sample treatment, e.g. chemical reactions, nor antibodies, and can be used for projective clinical studies of larger patient groups.