The purpose of the present study was to examine the influence of muscle group location and gender on the reliability of assessing the one-repetition maximum (1RM) test. Thirty healthy males (n = 15) and females (n = 15) who experienced at least 3 months of continuous resistance training during the last 2 years aged 18-35 years volunteered to participate in the study. The 1RM for the biceps curl, lat pull down, bench press, leg curl, hip flexion, triceps extension, shoulder press, low row, leg extension, hip extension, leg press and squat were measured twice by a trained professional using a standard published protocol. Biceps curl, lat pull down, bench press, leg curl, hip flexion, and squat 1RM's were measured on the first visit, then 48 hours later, subjects returned for their second visit. During their second visit, 1RM of triceps extension, shoulder press, low row, leg extension, hip extension, and leg press were measured. One week from the second visit, participants completed the 1 RM testing as previously done during the first and second visits. The third and fourth visits were separated by 48 hours as well. All four visits to the laboratory were at the same time of day. A high intraclass correlation coefficient (ICC > 0.91) was found for all exercises, independent of gender and muscle group size or location, however there was a significant interaction for muscle group location (upper body vs. lower body) in females (p < 0.027). In conclusion, a standardized 1RM testing protocol with a short warm-up and familiarization period is a reliable measurement to assess muscle strength changes regardless of muscle group location or gender.