This study represents the second validation phase of the Postpartum Worry Scale-Revised (PWS-R). As the PWS-R includes items tapping infant health and development concerns, we compare its psychometric properties with a sample of NICU mothers and the online sample used in the initial validation. We conduct a confirmatory factor analysis (CFA) to fit the latent factor structure previously validated with the online sample to the NICU sample. We also examine the reliability and construct validity of the PWS-R when used with a NICU sample. The PWS-R's reliability remains good and moderate concurrent correlations with theoretically similar constructs are shown with the newly created PWS-R factors. Model testing with the NICU sample reveals a different three factor structure than the four factor structure previously revealed with the online sample. The psychometric findings for the PWS-R provide continuing support for its use as a measure of postpartum worry; however, the differences in factor structure suggest that the PWS-R scores of high risk samples such as NICU parents should not be compared to those of community samples. Next steps in the iterative validation process and recommendations for use are considered, particularly with regard to high-risk samples.