Background <p>Critical congenital heart disease is a major cause of neonatal morbidity and mortality, requiring early detection and intervention. Pulse oximetry has been globally endorsed as a simple, cost-effective screening method. Despite its proven benefits, implementation in Egyptian neonatal care settings remains inconsistent. This study aimed to assess neonatal nurses’ knowledge, attitudes, and practices regarding the implementation of advanced newborn screening for Critical congenital heart disease using pulse oximetry across multiple hospitals in Egypt.</p> Methods <p>A descriptive, cross-sectional study was conducted from December 2024 to April 2025 in nine NICUs across five governorates. A convenience sample of 279 neonatal nurses was recruited. Data were collected using a structured questionnaire assessing sociodemographic characteristics, knowledge, attitudes, and observational practice checklists. Descriptive statistics, correlation analysis, and multiple linear regression were used for data analysis.</p> Results <p>The findings revealed unsatisfactory levels of knowledge (M = 3.56), negative attitudes (M = 12.55), and poor practice performance (M = 8.48). Strong positive correlations were found between knowledge, attitude, and practice (<i>r</i> = 0.59–0.68, <i>p</i> &lt; .001). Regression analysis identified total knowledge, total attitude, neonatal experience, training attendance, and bachelor’s education as significant predictors of better practice (<i>p</i> &lt; .01), while high nurse-to-patient ratios negatively impacted practice.</p> Conclusion <p>Neonatal nurses demonstrated low proficiency in Critical congenital heart disease screening, largely due to educational and institutional limitations. Structured training, standardized protocols, and improved staffing ratios are essential to enhance screening performance and ensure early Critical congenital heart disease detection.</p> Trial registration <p>Not applicable.</p>