Background In sub-Saharan Africa (SSA) 90 percent of babies acquired HIV/AIDS from infected mothers. Maternal knowledge about mother to child transmission (MTCT) of HIV/AIDS and its prevention is a cornerstone for elimination of MTCT of HIV/AIDS. Despite this, there is limited evidence about knowledge about MTCT of HIV/AIDS and its prevention and associated factors in SSA. Therefore, this study aimed to assess knowledge of MTCT of HIV/AIDS, its prevention (PMTCT) and, associated factors among reproductive-age women in sub-Saharan Africa. Objective To assess Knowledge about mother to child transmission of HIV/AIDS and its prevention and associated factors among reproductive-age women in Sub-Saharan Africa. Methods The recent SSA countries’ Demographic and Health Surveys (DHS), which were conducted from 2008/09 to 2018/19, was our data source. We appended 33 countries’ DHS data for our analysis. For our study, a total weighted sample of 350,888 reproductive-age women was used. Due to the hierarchical nature of the DHS data, we conducted a multilevel analysis. Finally, the adjusted odds ratio with its 95% confidence interval was reported, and variables with p-value≤0.05 were considered as significant predictors of knowledge of MTCT of HIV/AIDS and its prevention. Results In this study, 56.21% (95% CI: 56.05–56.38) of respondents had correct knowledge about MTCT of HIV/AIDS and its prevention among reproductive-age women in SSA. In the multilevel logistic regression analysis: being in the older age group, better education level, being from a rich household, having mass media exposure, having parity of one and above were associated with higher odds of knowledge about MTCT of HIV/AIDS and its prevention. However, being perceiving distance from the health facility as a big problem was associated with lower odds of knowledge about MTCT of HIV/AIDS and its prevention. Conclusion Knowledge about MTCT of HIV/AIDS and its prevention among reproductive-age women in SSA was low. Therefore, it is better to consider the high-risk groups during the intervention to increase awareness about this essential public health issue and to tackle its devastating outcome.