Abstract Introduction Timely identification of breast cancer is critical for successful treatment and prevention, reducing its burden. Despite its significance, breast cancer screening services in Ghana are underutilized, with considerable regional variations in uptake rates. We examined the regional differences in breast cancer screening uptake and its associated factors in Ghana. Methods We analyzed data from 15,014 women of reproductive age from the 2022 Ghana Demographic and Health Survey. Spatial mapping illustrated regional variations in screening uptake. Multilevel logistic regression was employed to examine the factors associated with breast cancer screening uptake. Results National breast cancer screening prevalence was 18.4%, with regional variations observed across the 16 regions. Eastern, Western, Brong Ahafo, and Greater Accra had the highest screening uptake. The likelihood of breast cancer screening was higher among women aged 45-49 (aOR = 3.59; 95% CI = 12.52, 5.12), Married women (aOR = 1.44; 95% CI = 1.17-1.77), Higher education, employment, health insurance, recent health facility visits, media access, and wealth index (middle: aOR = 1.45, 95% CI = 1.08, 1.94; richest: aOR = 1.91, 95% CI = 1.36, 2.70). Islamic and African Traditional affiliations decrease screening likelihood (aOR = 0.77; 95% CI = 0.63, 0.95; aOR = 0.46; 95% CI = 0.22, 0.97). Rural residence and specific regions (Ashanti, Oti, Savannah, Upper West) reduce odds, while living in North East increases odds (aOR: 1.86; 95% CI = 1.14-3.03). Conclusions Cost-effective breast cancer screening services are crucial, particularly in rural areas with geographical disparities. Culturally and religiously sensitive protocols are needed to address regional and religious differences. Expanding Ghana’s national health insurance to cover preventive measures like breast cancer screening is essential. Promoting awareness through conventional and digital platforms is crucial for advocating screening. Key messages • Addressing geographical and cultural differences crucial for equitable breast cancer screening in Ghana. • Expanding health insurance and raising awareness vital to promote screening access and reduce disparities.