Automated classification and visualization of fluorescent live cell microscopy images

Robotic, high-throughput microscopy is a powerful tool for small molecule screening and classifying cell phenotype, proteomic and genomic data. An important hurdle in the field is the automated classification and visualization of results collected from a data set of tens of thousands of images. We present a method that approaches these problems from the perspective of flow cytometry with supporting open-source code. Image analysis software was created that allowed high-throughput microscopy data to be analysed in a similar manner as flow cytometry. Each cell on an image is considered an object and a series of gates similar to flow cytometry is used to classify and quantify the properties of cells including size and level of fluorescent intensity. This method is released with open-source software and code that demonstrates the method's implementation. Accuracy of the software was determined by measuring the levels of apoptosis in a primary murine myoblast cell line after exposure to staurosporine and comparing these results to flow cytometry.