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Beazer, JD, Ljunggren, S, Karlsson, H, Davies, IG 
ORCID: 0000-0003-3722-8466 and Freeman, DJ
(2026)
A comparison of sequential density ultracentrifugation and density gradient ultracentrifugation for the proteomic analysis of high-density lipoprotein.
Frontiers in Cardiovascular Medicine, 13.
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A comparison of sequential density ultracentriguation and density gradient ultracentrifugation for the proteomic anlaysis of high-denisty lipoprotein.pdf - Published Version Available under License Creative Commons Attribution. Download (1MB) | Preview |
Abstract
Introduction: With the failure of high-density lipoprotein (HDL) cholesterol raising therapies to reduce cardiovascular risk, attention has turned towards HDL composition and function. There are several techniques for the isolation of lipoproteins from plasma, with density ultracentrifugation considered the gold-standard method. It is not known which form of density ultracentrifugation is optimal for HDL composition and function analyses.Methods: This study compared HDL composition by nLC-MS/MS and ELISA, subclass distribution by gel electrophoresis, and in vitro vascular anti-inflammatory function in HDL fractions isolated by sodium bromide sequential density ultracentrifugation (SDU) and iodixanol density gradient ultracentrifugation (DGU).Results: HDL composition differed between the two isolation techniques, with DGU-isolated HDL fractions containing a higher total protein content than SDU (10.14 ± 1.49 mg/mL compared to 3.18 ± 1.10 mg/mL respectively, p < 0.001, mean ± SD) but lower proteomic detection of key HDL proteins such as apolipoprotein A-I, apolipoprotein A-II and paraoxonase-1 in DGU-isolated HDL. HDL subclass distribution could not be determined in DGU HDL fractions due to contaminating plasma proteins. Vascular anti-inflammatory function was higher in DGU HDL (77.5 ± 4.4 % compared to 58.3 ± 4.8%, mean ± SD, p = 0.014) possibly due to the presence of confounding plasma proteins.Conclusion: Further steps may need to be added to DGU methods to obtain a ‘cleaner’ HDL fraction and therefore currently SDU is better suited to studies of HDL composition and function.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | 3201 Cardiovascular medicine and haematology |
| Subjects: | R Medicine > RC Internal medicine > RC1200 Sports Medicine |
| Divisions: | Sport and Exercise Sciences |
| Publisher: | Frontiers Media |
| Date of acceptance: | 23 April 2026 |
| Date of first compliant Open Access: | 18 May 2026 |
| Date Deposited: | 18 May 2026 15:13 |
| Last Modified: | 18 May 2026 15:13 |
| DOI or ID number: | 10.3389/fcvm.2026.1817349 |
| URI: | https://researchonline.ljmu.ac.uk/id/eprint/28593 |
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