title: T Cell Dynamic Activation and Functional Analysis in Nanoliter Droplet Microarray.
abstract: OBJECTIVE
  Characterization of the heterogeneity in immune reactions requires assessing dynamic single cell responses as well as interactions between the various immune cell subsets. Maturation and activation of effector cells is regulated by cell contact-dependent and soluble factor-mediated paracrine signalling. Currently there are few methods available that allow dynamic investigation of both processes simultaneously without physically constraining non-adherent cells and eliminating crosstalk from neighboring cell pairs. We describe here a microfluidic droplet microarray platform that permits rapid functional analysis of single cell responses and co-encapsulation of heterotypic cell pairs, thereby allowing us to evaluate the dynamic activation state of primary T cells.
  
  
  METHODS
  The microfluidic droplet platform enables generation and docking of monodisperse nanoliter volume (0.523 nl) droplets, with the capacity of monitoring a thousand droplets per experiment. Single human T cells were encapsulated in droplets and stimulated on-chip with the calcium ionophore ionomycin. T cells were also co-encapsulated with dendritic cells activated by ovalbumin peptide, followed by dynamic calcium signal monitoring.
  
  
  RESULTS
  Ionomycin-stimulated cells depicted fluctuation in calcium signalling compared to control. Both cell populations demonstrated marked heterogeneity in responses. Calcium signalling was observed in T cells immediately following contact with DCs, suggesting an early activation signal. T cells further showed non-contact mediated increase in calcium level, although this response was delayed compared to contact-mediated signals.
  
  
  CONCLUSIONS
  Our results suggest that this nanoliter droplet array-based microfluidic platform is a promising technique for assessment of heterogeneity in various types of cellular responses, detection of early/delayed signalling events and live cell phenotyping of immune cells.
authors.affiliation: Department of Pharmaceutical Sciences, Northeastern University, 360 Huntington Avenue, Boston, 02115 MA, USA.
authors.forename: Saheli
authors.initials: S
authors.lastname: Sarkar
authors.affiliation: Department of Pharmaceutical Sciences, Northeastern University, 360 Huntington Avenue, Boston, 02115 MA, USA.
authors.forename: Vinny
authors.initials: V
authors.lastname: Motwani
authors.affiliation: Department of Pharmaceutical Sciences, Northeastern University, 360 Huntington Avenue, Boston, 02115 MA, USA.
authors.forename: Pooja
authors.initials: P
authors.lastname: Sabhachandani
authors.affiliation: Department of Pharmaceutical Sciences, Northeastern University, 360 Huntington Avenue, Boston, 02115 MA, USA.
authors.forename: Noa
authors.initials: N
authors.lastname: Cohen
authors.affiliation: Department of Pharmaceutical Sciences, Northeastern University, 360 Huntington Avenue, Boston, 02115 MA, USA.
authors.forename: Tania
authors.initials: T
authors.lastname: Konry
keywords: Calcium; Dynamics; Heterogeneity; Immune response; Lymphocytes; Microfluidics; Single cell analysis; Time-lapse microscopy