Tracking Tau in Neurons: How to Transfect and Track Exogenous Tau in Primary Neurons.

Abstract

Primary murine neurons have proved to be an essential tool for the general investigation of neuronal polarity, polarized Tau distribution, and Tau-based neuronal dysfunction in disease paradigms. However, mature primary neurons are notoriously difficult to transfect with non-viral approaches and are very sensitive to cytoskeletal manipulation and imaging. Furthermore, standard non-viral transfection techniques require the use of a supportive glial monolayer or high-density cultures, both of which interfere with microscopy. Here we provide a simple non-viral liposome-based transfection method that enables transfection of Tau in low levels comparable to endogenous Tau. This allows the investigation of, for example, distribution and trafficking of Tau, without affecting other cytoskeleton-based parameters such as microtubule density or microtubule-based transport. Using this protocol, we achieve a profound transfection efficiency but avoid high overexpression rates. Importantly, this transfection method can be applied to neurons at different ages and is also suitable for very old cultures (up to 18days in vitro). In addition, the protocol can be used in cultures without glial support and at suitable cell densities for microscopy-based single cell analysis. In sum, this protocol has proven a reliable tool suitable for most microscopy-based approaches in our laboratory.