Background: The working die system used in dental prosthesis fabrication impacts the accuracy of dental prosthesis. Conventional, virtual, and 3D printed dies are used in various clinical scenarios. The accuracy of 3D printed models for diagnosis has been extensively studied. There is a lack of research on the marginal fit of interim single crowns made with 3D printed dies compared to conventional and virtual dies. Aim of the study: to evaluate the marginal fit of interim crowns fabricated using conventional, virtual, and three-dimensional printed working Dies. Materials and methods: A total of 18 interim crowns were milled and divided into three groups according to the die system on which they were fabricated; group A (fabricated on conventional dies), group B (fabricated on virtual dies) and group C (fabricated on 3D printed dies). Six physical impressions using polyvinyl siloxane material were taken to create conventional dies (group A) for maxillary right first molar. Additionally, six optical scans were performed to fabricate virtual dies (group B) and printed dies (group C). Interim crowns were milled, and a triple scan protocol was used to evaluate their marginal fit. Results: The virtual group demonstrated significantly better marginal fit compared to the other groups (p < 0.001). However, there was no statistically significant difference between the 3D printed and conventional groups. Conclusion: virtual dies yielded the highest marginal fit, while the conventional and 3D printed dies produced comparable results. The marginal fit of all interim crowns fell within the clinically acceptable range of 30 to 141 µm.