Intraspecific variation in antheridiogen response has been reported in several fern species. Here, we characterized and compared dose-mediated antheridiogen response among six populations spanning the range of Onoclea sensibilis, a species widely used for antheridiogen assays. A crude aqueous filtrate (CAF) of antheridiogen APT was obtained from cultures of Pteridium aquilinum and introduced into mineral agar at six concentrations ranging from 0 (control) to 10-1. Response was initially characterized for one population using three criteria: percentage of male gametophytes, number of antheridia per gametophyte, and mean surface area of gametophytes. Percent male was consistently the best descriptor of the response profile and was used to compare responses among populations. In all populations, gametophytes exhibited little or no response at CAF concentrations 10-5 and lower; a saturated response, almost all gametophytes being male at 10-3 and higher; and an intermediate response, usually between 50% and 80% males at 10-4. With few exceptions, response levels and profiles were very consistent among populations. Data and methodology provided herein indicate that use of the percent male criterion can provide a reliable quantitative assay using Onoclea that can facilitate future comparative research in antheridiogens.