Background: A sterile surgical marking pen is commonly used during anterior cruciate ligament reconstruction (ACLR) to outline the proposed skin incision and then to mark the graft during preparation. Once in contact with the skin, the pen is a potential source of bacterial transmission and subsequent infections after ACLR. Purpose/Hypothesis: The purpose of this study was to assess whether the skin marking pen is a fomite for contamination during arthroscopic ACLR. We hypothesized that there would be a difference in the rate of culture-positive pens between control pens and the study pens used to delineate the proposed skin incision. Study Design: Controlled laboratory study. Methods: Twenty surgical marking pens were collected prospectively from patients undergoing ACLR over a 12-month period. All patients underwent standard preoperative sterile preparation and draping procedures. Proposed incisions were marked with a new sterile pen, and the pen tip was immediately sent for a 5-day inoculation in broth and agar. Negative controls (unopened new pen) and positive controls (used to mark the skin incisions preoperatively) were also cultured. Additionally, blank culture dishes were observed during the growth process. All pens were removed from the surgical field before incision, and new marking pens were used when needed during the procedure. Results: Three of the 20 study pens (15%) demonstrated positive growth. All 3 pens grew species of Staphylococcus. None of the negative controls demonstrated growth, 6 of the 12 positive controls showed growth, and none of the blank dishes exhibited growth. Conclusion: This study found a 15% rate of surgical marking pen contamination by Staphylococcus during ACLR. It is recommended that the skin marking pen not be used for any further steps of the surgical case and be discarded once used. Clinical Relevance: Infections after ACLR are rare but may result in significant morbidity, and all measures to reduce them should be pursued. Surgeons performing ACLR should dispose of the surgical marking pen after skin marking and before intraoperative use such as graft markup.