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Objective This cross-sectional online study examined musculoskeletal pain (MSP) prevalence among German computer workers and its association with working from home (WFH) conditions.Material and Methods The online survey was conducted between September 2023 and April 2024 (t1). Current pain and pain experienced before the pandemic in early 2020 (t0, assessed retrospectively) were evaluated using the Brief Pain Inventory for various localizations. Symptoms of anxiety and depression were assessed by PHQ-4. A score for the ergonomic remote workstation equipment (WFH score) was established based on information about seating, table, and electronic workplace equipment. The relationship between WFH and newly developed MSP and worsening of MSP since the pandemic was modeled using logistic regression analyses and adjusted by several confounders.Results Data from 1,064 employees with at least four hours of daily computer work were analyzed. A total of 968 participants had worked from home (WFH employees), with a higher percentage at t1 compared to t0 (87% vs. 39%). According to the ergonomic WFH score, 74% of WFH employees had ergonomically well-equipped workstations. The highest MSP prevalence was observed for the lower back (42%) and neck (40%) at t1. WFH employees had a slightly higher risk than office workers for worsening of existing MSP (e.g., neck pain OR 1.62; 95% CI 0.83-3.19). Poorer workstation setups, longer daily screen time, or lack of regular participation in occupational health screenings were also risk factors for MSP. Additionally, more severe symptoms of anxiety or depression were associated with a higher risk of MSP (e.g., worsening low back pain OR 1.20; 95% CI 1.11-1.29).Conclusion To prevent MSP, employers should ensure that employees have the knowledge and equipment needed to set up and adjust their workstations according to ergonomic recommendations. They should also encourage regular participation in appropriate occupational health screenings.