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Abstract Background Down syndrome is a common chromosomal abnormality and prenatal screening can inform parents of the risk of their baby having Down syndrome. Little research has examined how decisions regarding both Down syndrome screening as well as diagnosis are made among women who are currently pregnant and how their decisions are influenced by their social contexts, specifically family and social media, using mixed methods. The study was to test the validity and reliability of a scale that measures pregnant women’s attitudes and decision-making concerning prenatal Down syndrome screening and diagnosis in urban areas of Taiwan. Methods We developed an item pool based on a literature review and in-depth interviews with 30 pregnant women recruited at two district hospitals in urban areas. The item pool was reviewed by a panel of experts and then administered to 300 women who had been pregnant for less than 24 weeks and had not received the Down syndrome screening tests. We used item analysis and exploratory factor analysis to validate the scale and test its reliability. Results The initial item pool had 54 items. After the expert review, three items were deleted. After the item analysis, 16 additional items were deleted. Exploratory factor analysis of the remaining items revealed four factors labeled – “Attitudes towards Down syndrome and Screening Tests,” “Important others’ Attitudes towards Down Syndrome,” “Influence of Important Others on Decision-Making,” and “Influence of Social Media on Decision-Making” – and 16 of the remaining items had satisfactory loadings on those factors, explaining 72.0% of the total variance. The Cronbach’s α values of the dimensions ranged between 0.75 and 0.90, demonstrating satisfactory internal reliability. Conclusions The scale has satisfactory validity and reliability, and can be used to understand pregnant women’s attitudes and decision-making regarding Down syndrome screening and diagnosis, and to help design tailored consultations for pregnant women in clinical settings.