AtlasScope encompasses 325 single-cell sequencing libraries, comprising 165 newly generated libraries and 160 sourced from publicly available datasets. These datasets span 131 species, including 41 mammals, 44 fishes, 10 birds, 7 reptiles, 3 amphibians, and 26 invertebrates. For both in-house and publicly sourced FASTQ files, we utilised the Cell Ranger pipeline to align reads, correct sequencing errors, and produce high-quality data for downstream single-cell RNA analysis. For public data in the form of original expression matrices, we read them into AnnData objects using Scanpy. The collected RDS files were processed by extracting the original count matrices, converting them into Loom files, and storing the associated metadata. We then read these Loom files and metadata with Scanpy and converted them into H5AD format.