Protocol for multiplexed in situ sequencing in tissue sections as an image-based spatial transcriptomic method. This is the second iteration of In Situ Sequencing (HybISS: Hybridization based In Situ Sequencing) based on the principles published in Ke et al. Nature Methods, 2013 and more recently Qian et al. Nature Methods, 2019. Here we present a modified method in detection chemistry using sequencing by hybridization approach for combinatorial decoding. This results in a more robust method for detecting larger probe panels for a more high-throughput in situ sequencing method.