The paper introduces DroNc-seq, a massively parallel single nucleus RNA-seq method that combines the advantages of single nucleus RNA-seq (sNuc-seq) and Drop-seq to profile nuclei at low cost and high throughput. DroNc-seq is designed to handle archived or difficult-to-dissociate tissues, such as brain tissue, by profiling 39,111 nuclei from mouse and human archived brain samples. The method demonstrates sensitive, efficient, and unbiased classification of cell types, including rare cell types, and captures fine distinctions between closely related cell types. DroNc-seq is compared to other methods in terms of throughput, sensitivity, and expression profiles, showing similar or better performance. The study also assesses the utility of DroNc-seq on archived human tissue, successfully identifying cell types and subtypes, rare cells, expression signatures, and activated pathways. The results highlight the potential of DroNc-seq for systematic single nucleus analysis of complex tissues, aiding in the creation of comprehensive atlases of human tissues and clinical samples.The paper introduces DroNc-seq, a massively parallel single nucleus RNA-seq method that combines the advantages of single nucleus RNA-seq (sNuc-seq) and Drop-seq to profile nuclei at low cost and high throughput. DroNc-seq is designed to handle archived or difficult-to-dissociate tissues, such as brain tissue, by profiling 39,111 nuclei from mouse and human archived brain samples. The method demonstrates sensitive, efficient, and unbiased classification of cell types, including rare cell types, and captures fine distinctions between closely related cell types. DroNc-seq is compared to other methods in terms of throughput, sensitivity, and expression profiles, showing similar or better performance. The study also assesses the utility of DroNc-seq on archived human tissue, successfully identifying cell types and subtypes, rare cells, expression signatures, and activated pathways. The results highlight the potential of DroNc-seq for systematic single nucleus analysis of complex tissues, aiding in the creation of comprehensive atlases of human tissues and clinical samples.