Heparin-based anticoagulants have been widely used for the prevention and treatment of venous thrombotic diseases, as well as for anticoagulation during cardiopulmonary bypass and hemodialysis. However, excessive heparin usage brings serious bleeding risk, necessitating immediate reversal of their anticoagulant activity. Additionally, to prevent bleeding during surgery and restore hemostatic function post-cardiopulmonary bypass and hemodialysis, it is also crucial to reverse heparin's anticoagulant effects. Currently, protamine sulfate (PS) is the only clinically approved antidote for heparin. However, its effectiveness against low molecular weight heparin (LMWH) and fondaparinux sodium is limited. Moreover, PS has great potential to trigger fatal allergic reactions. Despite these concerns, no successful clinical substitutes for PS have been developed. In the current work, drawing inspiration from the mechanism by which PS efficiently reverses heparin, we modified the cationic liposome with cationic amino acids, arginine and lysine, to serve as a broad-spectrum antidote (CRKRK-Lipo) for heparin-based anticoagulants. This modification not only enhances their reversal efficiency but also reduces the overall surface charge, potentially improving their biocompatibility. In the tail bleeding and liver injury mouse models, CRKRK-Lipo demonstrated reversal efficiency comparable to PS for heparin and superior reversal efficiency for LMWH and fondaparinux sodium. Notably, CRKRK-Lipo exhibited a wider therapeutic dose window and did not exhibit severe cytotoxicity or immunogenicity, in contrast to PS. It is worth noting that cationic liposomes without polypeptide modification also displayed a significant heparin reversal effect. Our findings not only offer a potential alternative for PS but also broaden the application fields of cationic liposome. STATEMENT OF SIGNIFICANCE: This study introduces the cationic liposomes as a novel and effective alternative to protamine sulfate (PS) for the functional reversal of heparin-based anticoagulants. The results reveal that both CRKRK-modified cationic liposomes (CRKRK-Lipo) and unmodified cationic liposomes (Lipo) showed comparable reversal efficiency to PS for UFH and superior reversal efficiency for LMWH and fondaparinux sodium, with a wider therapeutic dose window and reduced toxicity. This work offers an alternative strategy for detoxifying heparin-based anticoagulants and expands the biomedical applications of cationic liposomes.