Background: Stage 3 grade C periodontitis (S3GCP) has always been a challenge for clinicians. However, it is proposed that the use of lasers in addition to periodontal therapy can result in a more efficient therapy outcome.Objective: The aim of this clinical study was to determine the effects of additional application of diode laser (DL, 810 nm +/- 5) on clinical and microbiological values during Modified Widman Flap (MWF) periodontal surgery in the S3GCP patients.Methods: A total of 18 patients were randomly assigned to the test site (MWF + activeDL) and the control site (MFW alone). Clinical parameters and microbial samples were taken preoperatively, and postoperatively at 6 weeks, 3 months, and 6 months. Visual analog scale (VAS), tissue edema (TE), tissue color (TC), and pain medication (PM) consumption, were evaluated postoperatively at 10th day.Results: All bacteria were significantly decreased at follow-up times compared with preoperative amounts in both therapy sites (p < 0.05). Bacterial amounts of Treponema denticola, Prevotella intermedia, Tannerella forsythia, and Aggregatibacter actinomycetemcomitans at 6 weeks, Porphyromonas gingivalis and T. denticola at 3 months, T. denticola and A. actinomycetemcomitans at 6 months were statistically lower in the test site than the control site. All clinical parameters significantly improved after MWF surgery at follow-up times compared with preoperative values in intragroup comparisons, but a significant difference was not detected in the intergroup comparison. No differences were found in terms of VAS, PM, TE, and only TC was statistically more pink in the test site than the control site.Conclusions: The present study findings suggest that the use of DL together with MWF may have positive effects in the therapy of S3GCP patients by reducing the microbial load. Clinical Trial.org: NCT05108727.