Measuring Antibody-Mediated Tau Fibril Uptake in Microglia by Flow Cytometry

Microglia are brain-resident phagocytic cells, considered to be the innate immune cells of the central nervous system. Microglia respond to both infectious pathogens in the brain and sterile cellular debris, including the proteinaceous aggregates that accumulate in the brains of patients with Alzheimer's disease (AD). Microtubule-associated protein Tau is an intracellular protein that self-aggregates into neurofibrillary tangles in Alzheimer's disease and many other neurodegenerative diseases. Ongoing clinical trials are testing whether therapeutic antibodies specific to Tau protein aggregates can reduce pathological protein deposition and improve the course of disease. Data suggest that Tau-specific antibodies act on extracellular Tau aggregates by promoting uptake into microglia cells and thus preventing its prion-like spread to unaffected neurons. Here we describe a protocol to test the effect of Tau-specific antibodies on Tau uptake into microglia by flow cytometry. Recombinant Tau protein is fibrillized in vitro and tagged with a fluorescent label. Then, fibrillized Tau is incubated with the antibody of interest and applied to microglial cells in culture. Uptake of Tau into microglia is then assessed by measuring fluorescence intensity by flow cytometry. With slight modifications, this assay can be used to test effects of many antibodies, various Tau protein compositions, and different microglial sources in a high-throughput format.