Background: Becoming a mother is one of the principal transitions in a woman's life. which brings along the responsibility of caring for and maintaining life. Physical and psychological healthiness is changed leaving profound impacts on the quality of mothers' lives, especially those of first-time mothers, and making this postnatal period, in a way, unbearable to them. So, the study aimed to determine the effect of awareness program regarding motherhood preparation on first-time mothers' knowledge, practices, and emotional status. Subjects and method: Design: Quasi-experimental research design pre-post-test was utilized to achieve this study. Setting: The study was conducted at the postnatal department, postnatal outpatient clinic at Sohag University Hospital, followed by a telephone interview at 2 weeks after birth. Subjects: A convenience sample of 100 postnatal women was enrolled. Four tools were used for data collection: Tool (I): Structured interview questionnaire, Tool (II): Mother's knowledge, regarding self-care and their newborn care during the post-partum period (pre and post-test format), Tool (III): Mother's reporting practice regarding self-care and their newborn care during post-partum period (pre and post-test format), and Tool (IV): Depression, Anxiety, and Stress Scale (DASS) (pre and post-test format). The results: the present study revealed that there were highly statistically significant differences between mothers' knowledge, reporting practice, and their depression, anxiety, and stress levels regarding motherhood preparation pre and post-implementation of the awareness program. Conclusions: Awareness program regarding motherhood preparation application has a positive effect on improving first-time mothers' Knowledge, Practices, and emotional status pre-intervention than post-intervention. The study recommended conducting an awareness program for first-time mothers during pregnancy to inform them about self-care and their newborn care during the postpartum period to reduce their depression, anxiety, and stress levels.