Table 1.
An overview of methods used to isolate high-density lipoprotein subspecies. HDL: high-density lipoprotein
| METHOD | SUBSPECIES | ADVANTAGES | DISADVANTAGES |
|---|---|---|---|
| Ultracentrifugation | HDL 1,2,3 HDL 2b,2a, 3A,3b, and 3c | Size and density separation | Overlap with density similar lipoproteins like Lp (a) Imparts ionic strength and sheer stress and uses high salt concentrations |
| Two-dimensional gel electrophoresis | Charge: pre-beta, alpha, and pre-alpha Size: pre-beta-1 HDL, pre-beta-2, alpha 4- HDL, alpha 3- HDL, alpha-2 HDL, alpha-1 HDL, pre-alpha | Charge and size separation | Particles are not recoverable from the gel for functional analyses |
| Nuclear magnetic resonance spectroscopy | Small, medium, large. Newer algorithms can detect seven subclasses of HDL | Size separation High throughput Can quantify a particle number and particle size | Assumes that a constant number of methyl groups on each HDL subclass is constant, although newer algorithms account for variation Potential interference from plasma proteins Collinearity among subclasses needs to be accounted for in the analysis |
| Ion mobility | Very large, large, medium, small, and very small subspecies | Size separation Can quantify a particle number | HDL is isolated by ultracentrifugation first |
| Gel filtration | Six HDL subclasses and four lipid poor subspecies | Size separation Ability to isolate low abundant HDL particles Recoverability of HDL subclasses to study function | Time intensive Coelution with other plasma proteins such as albumin and globulins |
Figure 1.
The relationship between size exclusion chromatography and nuclear magnetic resonance (NMR) spectroscopy. Plasma was separated by size exclusion chromatography. Choline-containing phospholipid was quantified in each fraction using a kit from Wako. Eluted fractions were also quantified by NMR spectroscopy at LabCorp (formerly LipoScience) using an optimized version (LP4) of NMR LipoProfile. The average size particles from each of the gel filtration fraction quantified by NMR is shown in the table to the right and are consistent with those obtained by NMR. LDL: low-density lipoprotein; VLDL: very low-density lipoprotein; HDL: high-density lipoprotein; FPLC: fast protein liquid chromatography