This study investigated the influence of two different recruitment strategies on the reach and effect of a web-based multiple tailored smoking cessation program. From May 2009 until June 2010, Dutch adult smokers were recruited via mass media or general practices. Those who completed the baseline questionnaire were followed up during 6 weeks (two follow-ups). Differences between the two samples were assessed regarding baseline characteristics, retention rates, quit attempts and 24-hour point prevalence abstinence. Smokers recruited via general practices (N = 409) were significantly lower educated, less addicted, more motivated to quit smoking and to maintain non-smoking, more often female and more often suffering from cardiovascular or respiratory diseases than mass media respondents (N = 1154). They showed higher retention rates and were more likely to report a quit attempt (64.3 versus 50.7%) and abstinence (43.3 versus 33.1%). More respondents could be recruited via mass media, while general practices respondents showed higher retention rates and were more successful in quitting smoking, though these effects became non-significant when controlling for experimental condition and baseline differences. The choice for a particular recruitment strategy appeared to determine the number and type of smokers recruited and might consequently influence the intervention’s potential public health impact.