Automated Phenotyping via Cell Auto Training (CAT) on the Cell DIVE Platform We present a method for automatic cell classification in tissue samples using an automated training set from multiplexed immunofluorescence images. The method utilizes multiple markers stained in situ on a single tissue section on a robust hyperplex immunofluorescence platform (Cell DIVExe2x84xa2, GE Healthcare) that provides multichannel images allowing analysis at single cellsubcellular levels. The cell classification method consists of two steps: first, an automated training set from every image is generated using markertocell staining information. This mimics how a pathologist would select samples from a very large cohort at the image level. In the second step, a probability model is inferred from the automated training set. The probabilistic model captures staining patterns in mutually exclusive cell types and builds a single probability model for the data cohort. We have evaluated the proposed approach to classify: i) immune cells in cancer and ii) brain cells in neurological degenerative diseased tissue with average accuracies above 95%.

Classification of human white blood cells using machine learning for stainfree imaging flow cytometry Imaging flow cytometry (IFC) produces up to 12 different informationrich images of single cells at a throughput of 5000 cells per second. Yet often, cell populations are still studied using manual gating, a technique that has several drawbacks. Firstly, it is hard to reproduce. Secondly, it is subjective and biased. And thirdly, it is timeconsuming for large experiments. Therefore, it would be advantageous to replace manual gating with an automated process, which could be based on stainfree measurements originating from the brightfield and darkfield image channels. To realise this potential, advanced data analysis methods are required, in particular, machine learning. Previous works have successfully tested this approach on cell cycle phase classification with both a classical machine learning approach based on manually engineered features, and a deep learning approach. In this work, we compare both approaches extensively on the complex problem of white blood cell classification. Four human whole blood samples were assayed on an ImageStreamX MK II imaging flow cytometer. Two samples were stained for the identification of 8 white blood cell types, while two other sample sets were stained for the identification of resting and active eosinophils. For both datasets, four machine learning classifiers were evaluated on stainfree imagery using stratified 5fold crossvalidation. On the white blood cell dataset the best obtained results were 0.776 and 0.697 balanced accuracy for classical machine learning and deep learning, respectively. On the eosinophil dataset this was 0.866 and 0.867 balanced accuracy. From the experiments we conclude that classifying distinct cell types based on only stainfree images is possible with these techniques. However, both approaches did not always succeed in making reliable cell subtype classifications. Also, depending on the cell type, we find that even though the deep learning approach requires less expert input, it performs on par with a classical approach.