We have recently demonstrated by western blotting that, at birth, neonates show deficiency in protein kinase C (PKC) expression in T cells which normalises following maturation, evident for several PKC isozymes. Besides explaining the basis for immunodeficiency of immaturity, we found that the levels of PKCζ at birth could predict the risk of developing allergy in children. Supplementation of women during pregnancy with omega 3 fats increased the expression of PKCζ and this was associated with a decreased risk of allergic diseases. To undertake more extensive studies on the relationship between other PKC isozymes levels and cytokine production in T cell subsets, we have attempted to validate flow cytometry assays for these PKC isozymes. Commercially available antibodies marketed for flow cytometry analysis were examined for specificity against PKCα, βI, βII, γ, δ, ε, η, θ, ζ, ι/λ and μ by western blot analysis, using human peripheral blood mononuclear cells. A panel of monoclonal antibodies were selected based on these specificity results and tested for their ability to detect the isozymes in the different leukocyte subsets by flow cytometry. The results show that all PKC isozymes except PKCη are found in human leukocytes. PKCη is expressed in CD4, CD8 and NK cells but is not expressed in B lymphocytes, monocytes and neutrophils. Neutrophils showed highest expression of PKCα and μ, while PKC-δ was highly expressed in monocytes. This PKC isozyme assay could be performed with whole blood. Murine cells showed almost similar patterns of expression for these PKC isozymes in splenic cells. Using this flow cytometry assay deficiencies of PKC isozymes could be identified in human cord blood T cell subsets in whole blood assays. The technique enabled the monitoring of PKC isozyme levels during cord blood CD4 and CD8 T cell maturation in a tissue culture model.