Background: In contrast to conventional T lymphocytes invariant natural killer T cells (iNKTs) recognize lipid-based antigens presented by the class I MHC homolog CD1d. Upon activation, iNKTs can affect immune responses by promoting the secretion of Th1, Th2 or Th17 immune regulatory cytokine patterns. So far, iNKTs have been identified as important players in different types of immune responses. However, the role of iNKTs in a food allergic reaction is not well understood.

Methods: An iNKT reporter system was engineered by introducing the human receptor into a human leukemic Jurkat T cell line carrying an NF-kB-driven fluorescent transcriptional reporter construct (Jkt-iNKT). Antigen presenting cells were generated by expression of human CD1d in the murine thymoma cell BW (BW-CD1d). Reporter induction (NF-kB-driven eGFP-expression) was measured by flow cytometry. The specificity and sensitivity of our system was compared with the murine DN32 hybridoma iNKT cell line based assays. Food-derived lipid extracts and separated lipid fractions were obtained by Folch extraction and preparative thin layer chromatography, respectively. Lipid samples were screened applying a plate bound assay with recombinant CD1d as well as co-culture assays with Jkt-iNKT and BW-CD1d cell lines.

Results: Jurkat cells stably expressing the human iNKT TCR receptor (Jkt-iNKT) were generated and shown to specifically react with iNKT antigens presented in the context of CD1d. Detection limit for well-known iNKT cells antigens (α-GalCer and OCH) were similar for Jkt-iNKT and DN32 cell lines. Different food-derived lipid fractions from hazelnut, walnut, sunflower, and buckwheat were separated and analyzed by TLC.

Conclusions: Our Jurkat-based iNKT cell reporter cell line proved to be a useful tool (regarding readout and feasibility) to study the activation capacity of lipid molecules to activate human iNKT receptors. In addition, our reporter system is faster and cheaper as compared to assays based on murine hybridoma iNKT cell lines.