Time-Resolved Study of Nanoparticle Induced Apoptosis Using Microfabricated Single Cell Arrays

Abstract

Cell fate decisions like apoptosis are heterogeneously implemented within a cell population and, consequently, the population response is recognized as sum of many individual dynamic events. Here, we report on the use of micro-patterned single-cell arrays for real-time tracking of nanoparticle-induced (NP) cell death in sets of thousands of cells in parallel. Annexin (pSIVA) and propidium iodide (PI), two fluorescent indicators of apoptosis, are simultaneously monitored after exposure to functionalized polystyrene (PS−NH2) nanobeads as a model system. We find that the distribution of Annexin onset times shifts to later times and broadens as a function of decreasing NP dose. We discuss the mean time-to-death as a function of dose, and show how the EC50 value depends both on dose and time of measurement. In addition, the correlations between the early and late apoptotic markers indicate a systematic shift from apoptotic towards necrotic cell death during the course of the experiment. Thus, our work demonstrates the potential of array-based single cell cytometry for kinetic analysis of signaling cascades in a high-throughput format.

Correlation of mRNA delivery timing and protein expression in lipid-based transfection

“…We believe that fluorescence of the Cy5 labeled mRNAs is unquenched during the time of uptake due to unpacking of the lipid-based carriers. Other studies also used multiple fluorescent signals in single cell time lapse measurement to exploit signal correlations (36,37). Our findings are in agreement with a study by Yasar et al showing that the kinetics of mRNA nanocarrier binding on the cell surface is the same for transfected and nontransfected cells without giving further insights on transfection efficiency (26).…”

Area and Geometry Dependence of Cell Migration in Asymmetric Two-State Micropatterns

“…Cells seeded on micropatterns spread and adapt to the shape and size of the adhesive area (24). Regular lattices of micropatterns have been used to capture and strain single cells for a multitude of studies, including spindle-orientation (25,26), the influence of cell shape on cell polarization (27)(28)(29)(30)(31), and to monitor cell responses over time (32,33). For the investigation of cell migration dynamics, micropatterned lanes have proven useful as they confine cell motion to one dimension (34)(35)(36)(37).…”

A high throughput imaging database of toxicological effects of nanomaterials tested on HepaRG cells

“…The reference MNM, PS-NH 2 , was selected for its known chemical and physical properties and behaviour in vivo 12,13 . A detailed discussion on the sequence of events leading to PS-NH 2 -induced death of astrocytoma cells can be found in the literature 13 .…”