The authors describe the development of SpCas9-HF1, a high-fidelity variant of the *Streptococcus pyogenes* Cas9 (SpCas9) nuclease designed to reduce off-target genome-wide mutations. SpCas9-HF1 retains on-target activities comparable to wild-type SpCas9, with >85% of single-guide RNAs (sgRNAs) tested in human cells. Notably, with sgRNAs targeting standard non-repetitive sequences, SpCas9-HF1 rendered all or nearly all off-target events undetectable by genome-wide break capture and targeted sequencing methods. Even for atypical, repetitive target sites, the majority of off-targets induced by SpCas9-HF1 were not detected. The study suggests a general strategy for optimizing the genome-wide specificity of other RNA-guided nucleases and highlights the potential of SpCas9-HF1 for research and therapeutic applications.The authors describe the development of SpCas9-HF1, a high-fidelity variant of the *Streptococcus pyogenes* Cas9 (SpCas9) nuclease designed to reduce off-target genome-wide mutations. SpCas9-HF1 retains on-target activities comparable to wild-type SpCas9, with >85% of single-guide RNAs (sgRNAs) tested in human cells. Notably, with sgRNAs targeting standard non-repetitive sequences, SpCas9-HF1 rendered all or nearly all off-target events undetectable by genome-wide break capture and targeted sequencing methods. Even for atypical, repetitive target sites, the majority of off-targets induced by SpCas9-HF1 were not detected. The study suggests a general strategy for optimizing the genome-wide specificity of other RNA-guided nucleases and highlights the potential of SpCas9-HF1 for research and therapeutic applications.