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Comparison of Preparative High Density Lipoprotein Separation Methods for Subsequent Omic Analysis
Author(s) -
Zhu Chenghao,
Sacchi Romina,
Rhodes Christopher,
Tan Bowen,
Muchena John K.,
Lebrilla Carlito B.,
Zivkovic Angela M.
Publication year - 2016
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.30.1_supplement.684.11
Subject(s) - ultracentrifuge , chromatography , chemistry , lipidome , lipoprotein , high density lipoprotein , proteome , glycome , lipidomics , biochemistry , cholesterol , glycan , glycoprotein
Multiple methods are used to separate high density lipoprotein (HDL) from plasma, including an array of density‐based ultracentrifugation methods, as well as dextran sulfate separation. Ultracentrifugation is the gold standard for lipoprotein isolation and has been used to assess HDL for decades. However, it is thought that the high sheer stress and high salt concentrations in density ultracentrifugation based methods may lead to the loss of specific molecular species in HDL. Moreover, none of the current methods have been optimized and validated for subsequent omic anaylsis. In this study, several HDL separation methods were optimized and validated, including multiple sequential flotation and gradient ultracentrifugation methods, and the dextran sulfate precipitation method. These methods were compared using a standard pooled HDL sample and analyses were completed in triplicate. Lipidomic, proteomic, and glycomic analyses were performed on the separated HDL using mass spectrometry. Our results show that there were differences between methods in the proteome, lipidome, and glycome of HDL indicating that the specific HDL isolation protocol influences the compositional analysis of HDL.