The Tabula Sapiens is a reference human cell atlas containing single-cell transcriptomic data from more than two dozen organs and tissues. Here, we report Tabula Sapiens 2.0, which includes data from nine new donors, doubles the number of cells, and adds four new tissues. These new data include four donors with multiple organs contributed, thus providing a unique dataset in which genetic background, age, and epigenetic effects are controlled for. We illustrate applications of the Tabula Sapiens data in areas as diverse as gene regulation and aging. The Tabula Sapiens contains transcripts from nearly all human transcription factors, thereby providing putative cell-type specificity for nearly every human transcription factor and insights into their potential regulatory roles. We also analyze the transcriptomes of cells expressing canonical genes relating to cellular senescence, revealing the heterogeneity and context-dependent nature of senescence and providing new hypotheses into senescence-associated pathways.