Background: The diffusion of lung cancer screening programs has increased the detection of both solid and ground-glass opacity (GGO) sub-centimetric lesions, leading to the necessity for histological diagnoses. A percutaneous CT-guided biopsy may be challenging, thus making surgical excision a valid diagnostic alternative. CT-guided hydrogel plug deployment (BioSentry®) was recently proposed to simplify intraoperative nodule localization. Here, we report our initial experience. Methods: We evaluated 62 patients with single, small, peripheral, non-subpleural pulmonary GGO that was suspicious for cancer. All lesions were preoperatively marked, using CT-guidance, with a hydrogel plug (BioSentry®). Then, a uniportal video-assisted thoracoscopy (uniVATS) wedge resection was performed. If cancer was confirmed at the frozen section, a major lung resection was then performed. The study’s end points were the rates of intraoperative localization and of successful resection. Results: The hydrogel plug was correctly placed in 54 of the 62 cases, leading to an effective resection of the target lesion. In the remaining eight cases, the plug was displaced, and so the identification of pleural erosions due to the previous percutaneous procedure guided the resection. The uniVATS resection success rate was 98.3%. Conclusions: CT-guided hydrogel plug placement allowed for the successful detection of lung GGOs and resection with the uniVATS approach. This device allowed us to obtain lung cancer diagnoses and successfully treat 85.4% of cases.