The room temperature positron annihilation lifetime spectra and ⁵⁷Fe Mössbauer spectra were measured for pure Fe as well as for iron-based Fe_1−xRe_x, Fe_1−xOs_x, Fe_1−xMo_x, and Fe_1−xCr_x solid solutions, where x is in the range between 0.01 and 0.05. The measurements were performed in order to check if the known from the literature, theoretical calculations on the interactions between vacancies and solute atoms in iron can be supported by the experimental data. The vacancies were created during formation and further mechanical processing of the iron systems under consideration so the spectra mentioned above were collected at least twice for each studied sample synthesized in an arc furnace— after cold rolling to the thickness of about 40 μm as well as after subsequent annealing at 1270 K for 2 h. It was found that only in Fe and the Fe-Cr system the isolated vacancies thermally generated at high temperatures are not observed at the room temperature and cold rolling of the materials leads to creation of another type of vacancies which were associated with edge dislocations. In the case of other cold-rolled systems, positrons detect vacancies of two types mentioned above and Mössbauer nuclei “see” the vacancies mainly in the vicinity of non-iron atoms. This speaks in favour of the suggestion that in iron matrix the solute atoms of Os, Re, and Mo interact attractively with vacancies as it is predicted by theoretical computations and the energy of the interaction is large enough for existing the pairs vacancy-solute atom at the room temperature. On the other hand, the corresponding interaction for Cr atoms is either repulsive or attractive but smaller than that for Os, Re, and Mo atoms. The latter is in agreement with the theoretical calculations.