Objective: The aim of this study is to investigate the influence of local anesthetic (LA), operator experience level and needle type on patient procedural pain in relation to diagnostic lumbar puncture (LP). Methods: LP was performed with either a 22 gauge traumatic needle (22 TN) or a 22 gauge atraumatic needle (22 ATN). Immediately after LP patients documented a procedural pain score (PPS) on a 10-point Likert scale. Use of LA, needle type, anesthetic time interval (ATI), number of needle insertions and the LP operator experience level were registered. ATI was defined as the time from administration of LA to first needle insertion. Results: 104 patients had the LP procedure performed by 66 physicians (40 novices and 26 experienced physicians). Patients having the procedure performed by novices had a lower PPS of 2.56 if LA was administered compared to a higher PPS of 5.80 if LA was not administered (P = .046). Among experienced physicians there was no difference in PPS regardless of administration of LA. If novices administered LA, patient PPS was equal to patients having the procedure performed by an experienced operator. If novices performed the procedure with a 22 TN PPS decreased with increasing ATI (P = .01). No similar correlation was identified with the 22 ATN. Conclusion: Our study suggests that LP operator experience level, the needle type used and ATI may influence patient PPS. Further studies are necessary for final conclusions. These studies must consider these factors to avoid fault conclusions.