Abstract

Total Internal Reflection Fluorescence for Single Cell Analysis at the Level of Single Molecules

Alexander Asanov, Chief Scientific Officer, TIRF Labs Inc

Total Internal Reflection Fluorescence (TIRF) has become a method of choice for single molecule detection and is establishing itself as a powerful tool for other areas of lifesciences, including membrane biophysics, real-time microarrays, and molecular diagnostics.  Here, we report on a novel application of TIRF for single cell analysis. We combined TIRF with dielectrophoresis and real-time microarrays to perform lysis or electroporation of cells directly at the TIRF surface. Real-time microarrays printed at the TIRF surface allow for the multiplexed analysis of protein and nucleic acid markers obtained from single cells with the limit of detection down to single molecules. In this presentation, we will also describe the requirements for single molecule detection, and will compare prism-, lightguide-, and objective-based TIRF geometries for single molecule studies. This presentation will discuss the advantages and limitations of different TIRF geometries for single cell analysis, ion channel and dynamics of lipid rafts studies, and molecular diagnostics applications.