Oral Presentation Australasian Society for Immunology Annual Scientific Meeting 2014

Scrutinizing the magnitude and rhythm of ligand-induced intracellular calcium flux in T lymphocytes to deduce the strength of stimulus (#14)

Susan N Christo 1 2 , Kerrilyn R Diener 1 2 3 , Robert E Nordon 4 , Farid C Christo 5 , John D Hayball 1 2 6
  1. Sansom Institute, University of South Australia, Adelaide, SA, Australia
  2. Experimental Therapeutics Laboratory, Hanson Institute, Adelaide, SA, Australia
  3. School of Paediatrics and Reproductive Health, University of Adelaide, Adelaide, SA, Australia
  4. Graduate School of Biomedical Engineering, University of New South Wales, Kensington, NSW, Australia
  5. Barbara Hardy Institute, University of South Australia, Adelaide, SA, Australia
  6. School of Medicine, University of Adelaide, Adelaide, SA, Australia

Upon the interaction between the T cell receptor (TCR) and its cognate ligand, activated T cells elicit powerful cytolytic responses and can destroy pathogen-infected host cells. Understanding the parameters that result in optimal T cell activation may have applications in modulating T cells to the desired responses for immunotherapy regimens.

We have developed a method for capturing and activating individual T cells on functionalised material surfaces, enabling real-time individual and population-based analyses of calcium flux. When activated with anti-CD3 and variants of different TCR affinity, we find that there are distinct patterns in the magnitude and rhythm of calcium flux oscillations directly relating to the strength of stimulus. The rate of calcium flux was consistently higher in stronger bivalent antibody stimulation compared to its monovalent Fab counterpart. Interestingly, the addition of anti-CD28 co-stimulation reduced oscillatory potential but demonstrated a sustained calcium flux. This is consistent with the notion that calcium oscillations are the mechanism by which T cell can respond to lower affinity ligands, allowing transient calcium increases to cross intracellular thresholds that direct downstream events.  

These findings reveal a new and convenient method to evaluate T cell activation in real time and at a single cell level. The imaging process does not rely on complex microscopy and the population-based image analyses can be performed using simple open source programming. Our current investigation aims to make correlations between calcium flux patterns and T cell functional outputs at a single cell level.