The vibration performance of pedestrian structures attracts the attention of several studies, especially with respect to unfavorable operational conditions or possible damage scenarios. Given a pedestrian system, specific vibration comfort levels must be satisfied in addition to basic safety requirements, depending on the class of use, the structural typology and the materials. To this aim, guideline documents of the literature offer simplified single-degree-of-freedom (SDOF) approaches to estimate the maximum expected vibrations and to verify the required comfort limits. Most of these documents, however, are specifically calibrated for specific scenarios/structural typologies. Dedicated methods of design and analysis, in this regard, may be required for structural glass pedestrian systems, due to their intrinsic features (small thickness-to-size ratios, high flexibility, type and number of supports, live-to-dead load ratios, use of materials that are susceptible to mechanical degradation with time/temperature/humidity, etc.). Careful consideration could be then needed not only at the design stage, but also during the service life of a given glass walkway. In this paper, the dynamic performance of an in-service glass walkway is taken into account and explored via field vibration experiments. A set of walking configurations of technical interest is considered, involving 20 volunteers and several movement features. The vibration comfort of the structure is then assessed based on experimental estimates and existing guideline documents. The intrinsic uncertainties and limits of simplified approaches of literature are discussed, with respect to the performance of the examined glass walkway. In conclusion, the test predictions are also used to derive “perception index” data and scales that could support a reliable vibration comfort assessment of in-service pedestrian glass structures.