Effects of blood sample handling procedures on measurable inflammatory markers in plasma, serum and dried blood spot samples.

Author(s): Skogstrand K, Ekelund CK, Thorsen P, Vogel I, Jacobsson B, Nørgaard-Pedersen B, Hougaard DM

Publication: J Immunol Methods, 2008, Vol. 336, Page 78-84

PubMed ID: 18495149 PubMed Review Paper? No

Purpose of Paper

Conclusion of Paper

Studies

1. Study Purpose

   The purpose of this study was to determine the effects of storage of whole blood prior to centrifugation on 27 inflammatory markers in plasma and serum. After centrifugation, serum and plasma were stored at -20 degrees C until analysis.

   Summary of Findings:

   When whole blood specimens were stored at 4 degrees C for 4 h prior to centrifugation, interleukin (IL)-5, IL-17, Tumor necrosis factor beta (TNF-b), brain derived neurotrophic factor (BDNF), granulocyte-macrophage colony stimulating factor (GM-CSF), macrophage migration inhibitory factor (MIF), regulated upon activation normal T-cell expressed and secreted (RANTES), and C-reactive protein (CRP) were increased in plasma, and monocyte chemoattractant protein -1 (MCP-1), Macrophage inflammatory protein -1 alpha (MIP-1a) and -1 beta (MIP-1b), matrix metalloproteinase -9 (MMP-9), BDNF, soluble TNF Receptor 1 (sTNF R1), RANTES and CRP were increased in serum. After storage of whole blood for 4 h at room temperature prior to centrifugation, all analytes except IL-4, IL-6, IL-8, IL-10, IL-12 soluble IL-6 receptor alpha (sIL-6ra), TGF-beta and Neurothropin (NT)-3 were significantly increased in plasma and all analytes except IL-2, IL-4, IL-6, IL-8, IL-18, sIL-6ra, interferon (IFN)-gamma, Triggering receptor expressed in myeloid cells (TREM-1) and MIF-1 were increased in serum and after storage for 48 h at room temperature, only IL-4, IL-10, sIL-6ra and IFN-gamma were not significantly different in plasma and IL-2, IL-4, IL-5 and TGF-beta were not significantly different in serum than at baseline. Interestingly, after storage of blood for 4 h at 35 degrees C prior to centrifugation, the levels of 10 markers in plasma were significantly affected, but only IL-2, IL-4, IL-5, IL-12, IL-18, sIL-6ra and TREM-1 were not increased in serum. Generally the differences in inflammatory markers after storage of blood prior to centrifugation were larger for serum than plasma and when storage was at temperatures other than 4 degrees C.

   Biospecimens

   • Bodily Fluid - Plasma
   • Bodily Fluid - Serum
   • Bodily Fluid - Blood

   Preservative Types

   • None (Fresh)

   Diagnoses:

   • Not specified

   Platform:

   Analyte	Technology Platform
   Protein	Immunoassay

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Storage	Storage duration	0 h 4 h 24 h 48 h
   Storage	Storage temperature	4 degrees C Room temperature 35 degrees C
   Biospecimen Aliquots and Components	Blood and blood products	Plasma Serum
   Biospecimen Aliquots and Components	Centrifugation	Centrifugation delays investigated

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2. Study Purpose

   The purpose of this study was to determine the effects of storage temperature and duration on 25 inflammatory markers in DBS. After experimental storage, DBS were stored at -20 degrees C until analysis.

   Summary of Findings:

   Individual cytokines were differently affected by storage of dried blood spots prior to analysis, with both increases and decreases observed. IL-12, IL-18, TNF-beta, MIP-1a, MCP-1, BDNF and GM-CSF were the only cytokines generally affected by storage of DBS for 7 days at room temperature or 35 degrees C compared to DBS stored at -20 degrees C. When DBS were stored at 4 degrees C for 7 days, 1b, IL-6, IL-10, IL-18, IFN-gamma, TNF-beta, MIP-1b, BDNF, GM-CSF, NT-4 and CRP were significantly different then when stored at -20 degrees C, but after 30 days at 4 degrees C, only IL-18 was increased, and only TNF-beta and BDNF were decreased, compared to baseline. Protecting the DBS against humidity during storage at 4 degrees C did not prevent the changes in cytokine levels that were observed.

   Biospecimens

   • Bodily Fluid - Blood

   Preservative Types

   • Other Preservative

   Diagnoses:

   • Not specified

   Platform:

   Analyte	Technology Platform
   Protein	Immunoassay

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Storage	Storage duration	0 h 1 day 2 days 3 days 7 days 30 days
   Storage	Storage conditions	Exposed to humidity Protected from humidity
   Storage	Storage temperature	4 degrees C Room temperature 35 degrees C

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