Adsorptive separation using narrow-micropore adsorbents has demonstrated the potential to separate hydrogen isotopes. In this work, we employed an isotope-responsive separation using cobalt formate. A D2-responsive third sorption step was revealed, and consequently, a noticeable difference was observed in the uptakes of D2 and H2. This may have resulted from the additional space created for D2 due to its dense packing, as DFT calculations revealed that cobalt formate possesses 2.26 kJ/mol higher binding strength for D2 than for H2. The exploitation of this D2-responsive third sorption step renders a promising separation performance, with a D2/H2 selectivity of up to 44 at 25 K/1 bar. Lastly, cobalt formate was synthesized on a gram scale here, which makes it a prospect for commercialization.