Investigation of the solubility and the potentials for purification of serum amyloid A (SAA) from equine acute phase serum – a pilot study

BMC Research Notes

Mads Kjelgaard-Hansen²  Stine Jacobsen³  Jens Christian Sørensen¹  Michelle B Christensen² 

[1]Department of Food Science, University of Copenhagen, Thorvaldsensvej 40, 5th floor, Frederiksberg C 1870, Denmark

[2]Department of Veterinary Clinical and Animal Sciences, University of Copenhagen, Groennegaardsvej 3, ground floor, Frederiksberg C 1870, Denmark

[3]Department of Large Animal Science, University of Copenhagen, Hoejbakkegaard Allè 5, Taastrup 2630, Denmark

关键词: Horse;    Serum amyloid A;    Biochemical properties;   

Others  :  1142932
DOI  :  10.1186/1756-0500-6-152

 received in 2012-12-07, accepted in 2013-04-04,  发布年份 2013

【 摘 要 】

Background

Serum amyloid A (SAA) is useful as a diagnostic marker of systemic inflammation in horses, but only heterologous assays based on non-equine calibration and standardization are available for measurements of equine SAA. More accurate measurements could be obtained using purified species-specific SAA in native conformation for assay calibration and standardization. Further knowledge about the biochemical properties of SAA would facilitate a future production of native species-specific calibration material Therefore, the aim of the study was an investigation of the solubility and potentials for purification of equine SAA based on biochemical properties.

Freeze dried equine acute phase serum was dissolved in 70% 2-propanol, 8 M urea, and milli-Q water, respectively. Supercritical fluid extraction (SFE), size-exclusive chromatography (FPLC-SEC), and preparative isoelectric focusing (IEF) were performed in the attempt to purify. Immunostaining of IEF blots were used for isoform-specific detection of SAA in the preparations and purity was assessed by silverstained SDS-PAGE.

Findings

SAA was soluble in 70% 2-propanol, 8 M urea and Milli-Q water. SAA was not separated in the lipophilic or ampipathic fractions following SFE. SAA was included in a FPLC-SEC-fraction of 237 kDa, despite the molecular weight known to be much smaller, suggesting binding to other serum constituents. SAA precipitated following separation of other serum proteins by preparative IEF.

Discussion

No effective purification of SAA was achieved in the present study, but findings important for future investigations were made. The study suggested that SAA is not exclusively hydrophobic, but appears less hydrophobic when interacting with other serum components. These results suggest more complex aspects of solubility than previously believed, and indicate potentials for purification of native SAA.

【 授权许可】

   
2013 Christensen et al.; licensee BioMed Central Ltd.

【 图 表 】

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