The impact of blood-processing time on the proteome of human peripheral blood mononuclear cells.
Author(s): Bonilauri B, Santos MDM, Camillo-Andrade AC, Bispo S, Nogueira FCS, Carvalho PC, Zanchin NIT, Fischer JSDG
Publication: Biochim Biophys Acta Proteins Proteom, 2020, Vol. 1869, Page 140581
PubMed ID: 33301959 PubMed Review Paper? No
Purpose of Paper
This paper compared the proteomic profile of peripheral blood mononuclear cells (PBMCs) processed immediately and those obtained after a 24 h room temperature processing delay.
Conclusion of Paper
Hierarchical clustering based on the proteomic profile clustered specimens based on the processing delay rather than by patient. Of the 3195 proteins identified, 245 were differentially abundant based on processing delay. The proteins identified to be upregulated after a processing delay include those involved in exocytosis, neutrophil degranulation, neutrophil activation, immunity, granulocyte activation, leukocyte degranulation, myeloid cells activation, and immunity pathways and downregulated proteins were those involved in exocytosis regulation, localization, vesicle-mediated transport, cell activation, secretion, and exportation.
Studies
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Study Purpose
This study compared the proteomic profile of PBMCs processed immediately and those obtained after a 24 h room temperature processing delay. Blood was collected from four healthy males into EDTA Vacutainer tubes and immediately separated into two aliquots. PBMCs were isolated by Ficoll-Paque density gradient centrifugation immediately or after 24 h at room temperature (22°C). Proteins were extracted from PBMC using RapiGest surfactant in triethylammonium bicarbonate and centrifuged at 20,000 × g for 20 min at 4°C. Total proteins were quantified using Qubit, reduced, trypsin digested, and labeled with Isobaric Tags for Relative and Absolute Quantitation (iTRAQ 8plex). Purified labeled peptides were quantified by hydrophilic interaction chromatography MS/MS.
Summary of Findings:
Hierarchical clustering based on the proteomic profile clustered specimens based on the processing delay rather than by patient. Of the 3195 proteins identified, 245 were differentially abundant (fold-change >1.5 and p<0.01) based on processing delay with 101 upregulated and 144 downregulated after the 24 h processing delay. The proteins identified to have higher expression in fresh specimens were found to include those in pathways involving exocytosis regulation, localization, vesicle-mediated transport, cell activation, secretion, and exportation. In contrast, the proteins found to have increased expression in specimens subjected to a 24 h processing delay included those involved in exocytosis, neutrophil degranulation, neutrophil activation, immunity, granulocyte activation, leukocyte degranulation, myeloid cells activation, and immunity pathways.
Biospecimens
Preservative Types
- None (Fresh)
Diagnoses:
- Normal
Platform:
Analyte Technology Platform Peptide LC-MS or LC-MS/MS Protein LC-MS or LC-MS/MS Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Storage Storage duration 0 h
24 h
Biospecimen Aliquots and Components Centrifugation Centrifugation delays investigated