Background: Manual handling of patients without using proper devices built for this purpose is the most important risk factor causing musculoskeletal disorders (MSDs) among nursing staff. Objectives: The aim of the present study was to determine the prevalence of MSDs, assess the risk of manual handling of patients, and identify the risk factors related to the development of such disorders among nursing staff. Methods: This cross-sectional study was conducted in 2015 and included 220 nurses from 24 wards who were working in Baqiyatallah hospital, Tehran, Iran; their duties mainly involved patient handling. The participants were selected based on the random stratified sampling method. In this study, the Nordic musculoskeletal questionnaire, demographic questionnaire, movimentazione and assistenza di pazienti ospedalizzati (MAPO), and patient transfer assessment instrument (PTAI) methods were applied as data collection tools. In addition, Spearman correlation coefficients and binary and multiple logistic regressions were used to determine the possible relationships between the variables. Results: The prevalence of MSDs among the participants over the past 12 months was found to be 79.5%. The results for the risk factors showed that the lower back, neck, shoulders, and upper back were the most affected areas of the body. According to the MAPO and PTAI, 60% and 40% of the wards, respectively, are at risk of MSDs. A significant relationship between MAPO and PTAI was found, with a coefficient correlation of 0.252. Based on the statistical analysis, positively significant relationships were found between body mass index, gender, nurse-to-bed ratio, final PTAI and MAPO indices, and MSDs. Conclusions: The prevalence of MSDs among the nurses was high, and the occurrence of such disorders was found to be related to the scores of MAPO and PTAI indices. Based on the results, these two indices can be used as appropriate tools to assess the risk of MSDs in patient handling by nursing staff. The risk of developing MSDs can be lowered through interventional programs, such as providing and equipping hospitals with the tools required for patient handling and educating staff on how to properly use such devices. In addition, increasing the nurse-to-bed ratio will lower the burden for the nurses in patient handling.