Abstract Background In light of the increasing demands in health care, a call has been made for the development of new strategies. One of these strategies is placing a higher emphasis on individuals, who are expected to better manage their own health and illness. mHealth applications could increase this self-management behaviour among older adults. However, it is crucial to know the intention to use mHealth of older adults before implementing these services. Even less is known regarding differences between genders on factors influencing this intention to use mHealth applications. Objective The aim of this study was to study the gender differences regarding the relationship between technology acceptance factors and the intention to use mHealth applications in the Dutch elderly population. Methods We conducted a quantitative cross-sectional study using questionnaires. The participants were 65 years or older, lived independently or in a senior living facility, without cognitive impairment. Logistic regression with interaction terms was done to determine gender differences in the relationship between the intention to use mHealth applications and technology acceptance factors. Results While we found that half of the studied population had intention to use medical applications (50.3%) a notable difference was observed within gender groups which showed more men had intention to use medical applications rather than women (59.4% vs. 43.4% respectively). Adjusted logistic regression analysis per factor on the male and female part of the study population respectively showed that the factors Perceived usefulness (OR 21,69 and 2,39, resp.), Perceived ease of use (OR 7,21 and 2,74), Attitude toward use (OR 24,61 and 4,94), Sense of control (OR 4,12 and 2,67), Personal innovativeness (OR 2,54 and 1,58), Self-perceived effectiveness (OR 3,21 and 2,34), Service availability (OR 4,38 and 2,51) and Facilitating circumstances (OR 3,04 and 2,18) had a statistically significant influence on intention to use in both models. Logistic regression with interaction terms showed that two of the technology acceptance factors differed statistically significant in their relationship with intention to use when comparing females to males, namely Perceived usefulness (OR 0,11) and Attitude toward use (OR 0.24). Both factors were more strongly associated with intention to use for men compared to women. Conclusion Policymakers and interventions aiming to stimulate the uptake of mHealth applications should acknowledge gender differences. Interventions based on improving the Perceived usefulness and Attitude toward use among female users could be a means to stimulate the full potential of medical applications and improve the uptake.