The objective of this systematic review was to summarise population-based methods (i.e. methods that used representative data from populations) for estimating the population size of men who have sex with men (MSM), a high-risk group for HIV and other sexually transmissible infections (STIs). Studies using population-based methods to estimate the number or percentage of MSM or gay men were included. Twenty-eight studies met the inclusion criteria. Seven studies used surveillance data, 18 studies used survey data, and six studies used census data. Sixteen studies were conducted in the US, five were conducted in European countries, two were conducted in Canada, three were conducted in Australia, one was conducted in Israel, and one was conducted in Kenya. MSM accounted for 0.03-6.5% of men among all studies, and ranged from 3.8% to 6.4% in the US, from 7000 to 39100 in Canada, from 0.03% to 6.5% in European countries, and from 127947 to 182624 in Australia. Studies using surveillance data obtained the highest estimates of the MSM population size, whereas those using survey data obtained the lowest estimates. Studies also estimated the MSM population size by dimensions of sexual orientation. In studies examining these dimensions, fewer people identified as MSM than reported experience with or attraction to other men. Selection bias, differences in recall periods and sampling, or stigma could affect the estimate. It is important to have an estimate of the number of MSM to calculate disease rates, plan HIV and STI prevention efforts, and to allocate resources for this group.