A platform design for long-term high-resolution imaging of T lymphocyte-APC interactions in health and aging

Authors

• S Chakraborty
• B Dura
• J Voldman
• MH Sung

Abstract

Nuclear factor-kappaB (NF-kappaB) was discovered as a protein which binds to an enhancer of the kappa light chain in B cells. NF-kappaB can induce gene activity by binding discrete DNA elements in promoters or enhancers. The role of NF-kappaB is evolutionarily conserved in the immune system wherein it regulates the expression of effectors that further provide appropriate responses to pathogenic insults. We focus on discerning real-time NF-kappaB dynamics post activation of naïve CD4+ T cells, which is critical for our investigation of the functional significance of signaling dynamics during the initial antigen encounter in adaptive immunity. Elucidating the dynamic behavior of NF-kappaB involves long-term (24-48 hours) live imaging at a subcellular resolution. In addition to the general issues associated with any long-term imaging of live cells, a unique challenge arises from T cell motility. Microfluidics provides us the system where single cells can be trapped and maintained for a long duration microscopy. The use of microfluidics in imaging T cells for shorter time periods has been previously demonstrated. We wish to adapt existing tools to image T cells up to a few days. A fusion of microfluidics and long-term high-resolution imaging system will increase the throughput of quantitative live cell microscopy analysis for primary T lymphocytes. After we characterize the ‘proper’ dynamics in naïve T cell responses from young mice, we plan to apply our analysis to cells from old mice and determine features that deteriorate with age.

Scientific Focus Area: Immunology

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