Summary Maintenance of pluripotency is regulated by a network of transcription factors coordinated by Oct4, Sox2, and Nanog (OSN), yet a systematic investigation of the composition and dynamics of the OSN protein network specifically on chromatin is still missing. Here we have developed a method combining ChIP with selective isolation of chromatin-associated proteins (SICAP) followed by mass spectrometry to identify chromatin-bound partners of a protein of interest. ChIP-SICAP in mouse embryonic stem cells (ESCs) identified over 400 proteins associating with OSN, including several whose interaction depends on the pluripotent state. Trim24, a previously unrecognized protein in the network, converges with OSN on multiple enhancers and suppresses the expression of developmental genes while activating cell cycle genes. Consistently, Trim24 significantly improved efficiency of cellular reprogramming, demonstrating its direct functionality in establishing pluripotency. Collectively, ChIP-SICAP provides a powerful tool to decode chromatin protein composition, further enhanced by its integrative capacity to perform ChIP-seq., Graphical Abstract, Highlights • ChIP-SICAP isolates and identifies proteins that colocalize on chromatin • Chromatin composition around Oct4, Sox2, and Nanog depends on the pluripotent state • Trim24 is part of the pluripotency network and promotes reprogramming • ChIP-SICAP allows recovery of DNA for sequencing with ChIP-seq quality, Rafiee et al. introduce ChIP-SICAP as a novel method to identify chromatin-bound proteins that colocalize with a bait protein on DNA. Applied to Oct4, Sox2, and Nanog in ESCs, they identify Trim24 as a novel protein that is functionally involved in the pluripotency network by promoting cellular reprogramming.