Background Except for one study in 2004, the literature has no data on the information‐seeking behaviour of lesbian, gay, bisexual, transgender and queer/questioning (LGBTQ) health professionals. After a decade of change for LGBTQ people, and the growth of electronic information sources and social networks, it is appropriate to revisit this subject. Objectives To gain an updated understanding of the information‐seeking behaviour of LGBTQ health professionals and of how medical libraries can provide a culturally competent service to such users. Methods A mixed‐methods approach was adopted combining a Web‐based questionnaire with email follow‐up discussions. One hundred and twenty‐three complete responses were received, mostly from the USA and Canada, between November 2012 and October 2013. Results LGBTQ health professionals remain more comfortable seeking LGBTQ health information from a medical librarian whom they know to be LGBTQ because they perceive LGBTQ librarians as more likely to have specialist knowledge, or through concern that non‐LGBTQ librarians may be more likely to react in a stigmatising or discriminatory way. The study also provides evidence suggesting that online chat has marginal appeal for respondents seeking LGBTQ health information, despite its anonymity. Medical libraries seeking to demonstrate their cultural competency should provide visible evidence of this, such as through the creation of dedicated resource lists, promotion of LGBTQ literature on the library's website, and display of other symbols or statements supporting diversity. Conclusion Opportunities exist for LGBTQ health professionals and medical librarians to work together to ensure that medical libraries are culturally competent and welcoming spaces for LGBTQ patrons, that library collections match their needs, and in the creation of guides to ensure maximum access to the results of LGBTQ health research. Medical libraries should also consider nominating and, if necessary, training a specialist in LGBTQ health information. Such measures are more likely to be successful than reliance on online chat, despite contrary suggestions in the literature.