Cancer Cell Line Authentication (CCLA) is a web server to authenticate human cancer cell lines (CCLs) using expression profiles from RNA-Seq or microarray data. In CCLA, we curated expression profiles and gene singtures of 1,291 human CCLs from various resources. CCLA employed the single sample gene set enrichment score and global pair-wise distance algorithm to automatically and accurately authenticate CCLs. By applying on comprehensive validation datasets 719 samples of 461 CCLs from 15 independent studies CCLA achieved an accuracy of 96.58% or 92.15% ), and the accuracy increased to 100% or 95.11% when considering results occurred within top3 outcomes for microarray or RNA-Seq data. Users can freely and conveniently authenticate CCLs using gene expression matrix or NCBI GEO accession on CCLA.