Objective To investigate the influence of different puncture needle types and ultrasonic characteristics of thyroid nodules on the sampling satisfaction rate of ultrasound-guided fine needle biopsy specimen. Methods A total of 185 patients with thyroid nodules (185 nodules)who underwent fine-needle aspiration biopsy in Nanjing Tongren Hospital from August 2020 to August 2022 were included. All nodules received 23G and 25G ultrasound guided fine-needle aspiration biopsy (US-FNAB). According to nodular ultrasonic characteristics (nodule size, coarse calcification, blood supply, echo), puncture specimens were classified into satisfactory and unsatisfactory categories according to The Bethesda system for reporting thyroid cytopathology (TBSRTC). The influence of puncture needle types and nodular ultrasonic characteristics on the sampling satisfaction rate were evaluated. Results There was no statistical difference in the overall sampling satisfaction rate between the 23G group and the 25G group (85.4% vs 88.1%, P=0.443). According to the maximum diameter, the nodules were divided into three groups. The satisfactory rate of specimen collection in the group ≤ 0.5cm was lower than that in the group 0.5 - 1.0cm and group ≥ 1.0cm (P<0.05). According to nodule calcification, the patients were divided into coarse calcification group and other groups. The satisfaction rate of 23G fine needle in the coarse calcification group was higher than that of 25G fine needle (76.5% vs 52.9%, P<0.05). There was no statistical difference in the satisfaction rate of the two kinds of fine needle in other groups. According to the blood supply, the nodules were divided into the blood supply rich group and the blood supply deficient group. The satisfaction rate of 25G fine needle in the blood supply rich group was higher than that of 23G fine needle (83.3% vs 61.9%, P<0.05). There was no statistical difference in the sample collection satisfaction rate between 23G and 25G fine needles in the blood supply deficient group. The nodules were divided into hypoechoic, isoechoic and hyperechoic groups according to their echo. There was no statistical difference in sampling satisfaction rate between 23G and 25G fine needles in different echo groups (P ≥ 0.05).Conclusion Small thyroid nodule (maximum diameter ≤ 0.5cm), accompanied by coarse calcification or rich blood supply will affect the sampling satisfaction rate.