PURPOSE: Studies using magnetic resonance imaging to assess lumbar multifidus cross-sectional area frequently utilize T1 or T2-weighted sequences, but seldom provide the rationale for their sequence choice. However, technical considerations between their acquisition protocols could impact on the ability to assess lumbar multifidus anatomy or its fat/muscle distinction. Our objectives were to examine the concurrent validity of lumbar multifidus morphology measures of T2 compared to T1-weighted sequences, and to assess the reliability of repeated lumbar multifidus measures. METHODS: The lumbar multifidus total cross-sectional area of 45 patients was measured bilaterally at L4 and L5, with histogram analysis determining the muscle/fat threshold values per muscle. Images were later re-randomized and re-assessed for intra-rater reliability. Matched images were visually rated for consistency of outlining between both image sequences. Bland-Altman bias, limits of agreement, and plots were calculated for differences in total cross-sectional area and percentage fat between and within sequences, and intra-rater reliability analysed. RESULTS: T1-weighted total cross-sectional area measures were systematically larger than T2 (0.2 cm2), with limits of agreement <±10% at both spinal levels. For percentage fat, no systematic bias occurred, but limits of agreement approached ±15%. Visually, muscle outlining was consistent between sequences, with substantial mismatches occurring in <5% of cases. Intra-rater reliability was excellent (ICC: 0.981-0.998); with bias and limits of agreement less than 1% and ±5%, respectively. CONCLUSION: Total cross-sectional area measures and outlining of muscle boundaries were consistent between sequences, and intra-rater reliability for total cross-sectional area and percentage fat was high indicating that either MRI sequence could be used interchangeably for this purpose. However, further studies comparing the accuracy of various methods for distinguishing fat from muscle are recommended.