The aim of this study was to explore and assess office-based ultrasound-guided intraosseous and intra-articular infiltrations of plasma rich in growth factors (PRGF) in patients with moderate and severe knee osteoarthritis (KOA). Seventy-nine patients (30 women and 49 men) with grade 3–4 KOA according to the Kellgren–Lawrence classification participated in the study. All patients were treated with a minimally invasive technique using local anesthesia WALANT (wide-awake local anesthesia no tourniquet) in the ambulatory setting. A PRGF intra-articular infiltration and two intraosseous infiltrations in the tibial plateau and femoral condyle were performed weekly for a total of three sessions. The evaluation of the results was carried out using knee injury and osteoarthritis outcome score (KOOS) at baseline and post-treatment. After a follow-up period of 11 months (median) [interquartile range, 7–14], all the KOOS domains showed statistically significant improvement (p< 0.001). Moreover, 88% of the patients showed a pain reduction of at least 10 points (minimally clinically important improvement) from pre- to post-treatment. Our retrospective study using the in-office procedure of ultrasound-guided combination of intra-articular and intraosseous infiltrations of PRGF is a safe and efficacious approach for the treatment of grade 3–4 knee osteoarthritis.