Multiplexed DNA fluorescence in situ hybridization (FISH) imaging technologies have been developed to map the folding of chromatin fibers at tens of nanometer and tens of kilobase resolution in single cells. However, computational methods to reliably identify chromatin loops from such imaging datasets are still lacking. Here we present a Single-Nucleus Analysis Pipeline for multiplexed DNA FISH (SnapFISH), to process the multiplexed DNA FISH data and identify chromatin loops. SnapFISH can identify known chromatin loops from mouse embryonic stem cells with high sensitivity and accuracy. In addition, SnapFISH obtained comparable results of chromatin loops across datasets generated from diverse imaging technologies.