A standardized method to study immune responses using porcine whole blood

Area of Science:

• Immunology
• Veterinary Medicine
• Assay Development

Background:

• Peripheral blood mononuclear cell (PBMC) isolation for in vitro immune assessment is time-consuming and variable.
• Whole blood assay (WBA) offers a faster, more physiological, and less resource-intensive alternative.

Purpose of the Study:

• To develop and validate a porcine whole blood assay (WBA) for in vitro immune response evaluation.
• To optimize WB dilution for immune cell stimulation and cytokine analysis.

Main Methods:

• Heparinized whole blood (WB) was diluted and stimulated with phorbol myristate acetate and ionomycin.
• Interferon-gamma (IFN-γ) secreting T-cells were quantified via flow cytometry.
• Tumor necrosis factor-alpha (TNF-α) and other cytokines were measured in supernatants after stimulation with various agents including LPS, poly I:C, Streptococcus suis, and PRRSV.

Main Results:

• Optimal immune cell response, indicated by IFN-γ+ T-cells and TNF-α levels, was observed at a 1/8 WB dilution.
• Lipopolysaccharide (LPS) and polyinosinic:polycytidylic acid (poly I:C) significantly increased TNF-α and IL-10 levels.
• Bacterial and viral stimuli induced specific cytokine profiles and altered immune cell populations.

Conclusions:

• The developed WBA provides a rapid, reliable, and simple method for assessing porcine immune responses.
• Dilution of WB is crucial for effectively triggering immune cells in this assay.