Single-cell cytokine profiles in normal humans: comparison of flow cytometric reagents and stimulation protocols.

Author(s): Jason J, Larned J

Publication: J Immunol Methods, 1997, Vol. 207, Page 13-22

PubMed ID: 9328582 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 using monocytes rather than whole blood, PHA stimulation, fluorochrome type and reagent source on the production of cytokines.

   Summary of Findings:

   Cytokine production was induced in the presence or absence of PHA. A higher proportion of CD8+ cells produced IFN than CD4+ cells, and a higher proportion of CD4+ cells produced IL-2 than CD8+ cells, but a similar proportion of CD4+ and CD8+ cells produced TNF-alpha. Generally, the addition of PHA to stimulate cytokine production did not have a great effect but still resulted in slightly increased percentages of CD4+ and CD8+ cells producing TNF alpha, slightly increased percentages of CD8+ cell producing IFN-gammma, and a decreased proportion of CD4+ and CD8+ cells producing IL-2. A smaller proportion of mononuclear cells produced IFN-gamma and IL-2 than whole blood cells, but a similar percentage of whole blood and monocyte cells produced TNF-alpha. The variability due to fluorochrome or reagent source was smaller than that between cell types or stimulation protocols, but in general the percentages of positive cells were lower using the PharMingen reagents than the Becton Dickinson (BD) reagents.

   Biospecimens

   • Bodily Fluid - Blood
   • Cell - Blood

   Preservative Types

   • None (Fresh)

   Diagnoses:

   • Normal

   Platform:

   Analyte	Technology Platform
   Cell count/volume	Flow cytometry
   Protein	Flow cytometry

   Pre-analytical Factors:

   Classification	Pre-analytical Factor	Value(s)
   Biospecimen Aliquots and Components	Blood and blood products	Mononuclear cells Whole blood
   Flow cytometry Specific	Reaction solution	PMA, PHA and ionomycin in RPMI-1640 buffer with BFA and 2 mM L-glutamine PMA and ionomycin in RPMI-1640buffer with BFA and 2 mM L-glutamine without PHA RPMI-1640buffer with BFA PharMingen BD
   Flow cytometry Specific	Detection method	Fluorescein Phycoerythrin

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