The analysis of heterogeneous subpopulations of circulating tumor cells (CTCs) is critical to enhance our understanding of cancer metastasis and enable non-invasive cancer diagnosis and monitoring. The phenotypic variability and plasticity of these cells – properties closely linked to their clinical behavior – demand techniques that isolate viable, discrete fractions of tumor cells for functional assays of their behavior and detailed analysis of biochemical properties. Here, we introduce the Prism Chip, a high-resolution immunomagnetic profiling and separation chip which harnesses a cobalt-based alloy to separate a flowing stream of nanoparticle-bound tumor cells with differential magnetic loading into ten discrete streams. Using this approach, we achieve exceptional purity (5.7 log white blood cell depletion) of isolated cells. We test the differential profiling function of the integrated device using prostate cancer blood samples from a mouse xenograft model. Using integrated graphene Hall sensors, we demonstrate concurrent automated profiling of single cells and CTC clusters that belong to distinct subpopulations based on protein surface expression.