CRISPR genome editing is a promising tool for translational research but can cause undesired editing outcomes, both on target at the edited locus and off target at other genomic loci. Here, we investigate the occurrence of deleterious on-target effects (OnTEs) in human stem cells after insertion of disease-related mutations by homology-directed repair (HDR) and gene editing using non-homologous end joining (NHEJ). We identify large, mono-allelic genomic deletions and loss-of-heterozygosity escaping standard quality controls in up to 40% of edited clones. To reliably detect such events, we describe simple, low-cost, and broadly applicable quantitative genotyping PCR (qgPCR) and single-nucleotide polymorphism (SNP) genotyping-based tools and suggest their usage as additional quality controls after editing. This will help to ensure the integrity of edited loci and increase the reliability of CRISPR editing. Display omitted •On-target effects (OnTEs) are present in up to 40% of human CRISPR-edited iPSC clones•OnTEs are frequently missed by standard quality controls, such as locus sequencing•Unnoticed OnTEs strongly affect phenotype formation in an iPSC Alzheimer model•Simple and broadly applicable qgPCR and SNP genotyping-based tools reliably detect OnTEs Weisheit et al. show that deleterious on-target effects, such as large, mono-allelic deletions or loss of heterozygosity, are widespread in human iPSCs after CRISPR/Cas9 editing both via HDR and NHEJ. They describe simple, low-cost, and broadly applicable quantitative genomic PCR (qgPCR) and SNP genotyping-based tools to reliably identify such on-target effects.