Background: In recent decades, many workers from rural areas in China migrated to urban cities in search of a better livelihood. Due to the household registration policy and other financial barriers, more than 40 million of children were left behind in their rural home by their migrated parents in 2015. In this cross-sectional study, we aimed to investigate the influence of being left behind on these children's physical and mental health. Methods: A self-administered questionnaire was completed by participants about their demographic background and health status. Chi-square Test was conducted to investigate the influence. Results: A total of 1662 participants responded and completed all the questions in the questionnaire. Significant differences existed between left-behind children group and non-left-behind children group on several health issues such as not going to school due to sickness (p = 0.080), completeness of the vaccination scheme (p = 0.036) and feeling of loneliness (p = 0.039). However, regarding symptoms like fever, cough or respiratory difficulties, diarrhea and twitch, as well as mental health problems like unhappiness and insomnia, no significant difference was found. Gender difference was also demonstrated showing that girls were more vulnerable than boys to certain symptoms and emotional problems. Conclusion: This study indicated that both being left-behind and gender had an impact on the children's health. It is necessary to further reform the household registration system to improve rights of equal access to employment, education and health resources for workers and their children from rural areas in China.