Purpose: Despite some spinal tuberculosis cases having typical imaging findings, histological/ microbiological evidence is required for definitive diagnosis and management. Percutaneous biopsy is suitable for histopathologic and bacteriologic evaluation of vertebral lesions, as it is a quick and minimally invasive technique to obtain specimens from deep structures and is advantageous for open surgical biopsy. Based on a minimally invasive technique, herein, we describe a newly designed percutaneous transpedicular biopsy system for lumbar vertebral lesion biopsy. Methods: Fifty-six patients who underwent percutaneous transpedicular vertebral biopsy under fluoroscopic guidance were evaluated. Biopsy specimens were obtained by passing a proprietary 6-mm diameter biopsy instrument through the pedicle and into the disease site under C-arm fluoroscopy. Specimens were sent for histological and bacteriologic analyses. Results: There were 36 males and 20 female patients of mean age 47.64 years (range 22-80 years). Biopsies were performed on 56 lumbar vertebral bodies. The average operative time were 15.6 minutes (range 13-18 minutes). All 56 patients (54 tubercular pathology, one metastasis, and one osteoporotic fracture with chronic nonspecific inflammation) had definitive histological/ microbiological diagnosis. Two patients (3.57% of 56 patients) developed a biopsy site small hematoma. Conclusions: A newly designed percutaneous transpedicular biopsy system was successfully used for lumbar vertebral biopsy under local anesthesia. It was easy to use and was safe with a low complication rate for definitive diagnosis of lumbar spinal tuberculosis and could be performed with minimal morbidity and high diagnostic yield as an outpatient procedure.