Motivation: Chromatin Immuno-Precipitation followed by Sequencing (ChIP-Seq)is used to determine the binding sites of any protein of interest. ChIP-Seq data suffer from being more qualitative than quantitative. The recent use of spike-in controls along with the standard protocol tackled this problem. However, no dedicated tool is available for a robust evaluation of this new ChIP-Seq approach. Results: We developed ChIPSeqSpike, an R/Bioconductor package that enables ChIP-Seq spike-in normalization, assessment and analysis. Ready to use scaled bigwig files and scaling factors values are obtained as output. ChIPSeqSpike also provides tools for ChIP-Seq spike-in assessment and analysis through a versatile collection of graphical functions. Availability: The package is implemented in R (as of version 3.4) and is available from Bioconductor at the URL: https://www.bioconductor.org/packages/3.7/bioc/html/ChIPSeqSpike.html, where installation and usage instructions can be found.