The exposure to air fine aerosols can cause health effects due to inhalations of alpha emitters such as 222Rn daughters. Lead-210 and 210Po are mainly associated to aerosols with median aerodynamic diameter lower than 1 μm. The 210Po is characterized by having a high radiotoxicity. The precise measurement of 210Po in surface air aerosols is usually quite complex due to the significant contribution of the 210Pb on their concentrations. Additionally, there is no possible means to manufacture a certified material to validate the measurements of 210Pb and 210Po concentrations in surface air aerosols. For these reasons, this study aims to develop a novel and comprehensive methodology to validate 210Po measurements in surface air aerosols by preparing in our laboratory “standard samples” with known activities of both 210Pb and 210Po. A detailed sensitivity analysis on the precision of 210Po concentration measurements has been carried out as a function of the involved variables such as sampling time, time elapsed between the sampling start and 210Po self-deposition and the 210Po/210Pb activity ratio in surface air aerosols. This study is necessary to find the optimum conditions for a precise measurement of 210Po in surface air aerosols. In addition, this methodology has been applied for the determination of 210Po concentrations in 30 samplings campaigns of atmospheric aerosols carried out at the El Carmen campus (Huelva province) from March 25th to July 15th, 2022. The results obtained for 210Pb and 210Po concentrations and atmospheric aerosol residence times (via 210Po/210Pb activity ratio) were consistent with other previous works.