Background As the public increasingly uses the internet to search for resources and information regarding health and medicine, it is important that health care organizations provide adequate web resources. Website usability refers to the ease of user experience on a website. In this study, we conducted usability analyses on digital health center websites. Objective The primary aims of this study were to (1) replicate a preexisting usability scoring methodology for digital health centers; (2) apply and test this replicated usability scoring methodology on a sample set of digital health center websites; and (3) derive recommendations from the results on potential areas of improvements for our sample of digital health center websites. Methods Website usability testing was conducted from March 1, 2020, to March 15, 2020. We replicated a methodology and scoring system from previous literature and applied them to digital health center websites. Our sample included 67 digital health centers that were affiliated with US universities or hospital systems. Usability was split into the following four broad categories: accessibility, marketing, content quality, and technology. Usability tools were used to score websites in each of the four categories. The composite of the key factors of each category was used to generate a general usability and overall usability score for each website. Results The category with the highest average score (6.3) was content quality. The content quality score also had the highest SD (2.18) and an SE of 0.27. The lowest performing category was technology, which had an average score of 0.9. The technology score also had the smallest SD (0.07) and an SE of 0.01. Conclusions Our data suggest that content quality, on average, was the highest scoring variable among digital health center websites. As content is crucial to digital health knowledge, it is justified that digital health centers invest more resources into creating quality content. The overall lowest scoring variable was technology. Potential reasons for this finding include designated funding for servers, a lack of regulatory frameworks for social media presence and liability, and infrequent website audits. An easy approach for improving this variable is increasing website speed. Accessibility is another area that organizations can potentially improve. We recommend that these organizations perform periodic audits of their web presence with usability tools.