The cell glycocalyx is an attractive model for surface modification of liposomes with the objectives of tissue targeting and prolonged circulation time. Here, we reported on glycocalyx-mimicking liposomes, prepared by incorporating a glycolipid of 3'-sulfo-Lewis a (SuLe(a))-PEG-DSPE with a headgroup of SuLe(a) and a spacer of poly(ethylene glycol) (PEG) linked to two hydrophobic tails. This PEG spaced structure is used to mimic the extended structure of P-selectin glycoprotein ligand 1 (PSGL-1) on activated leukocytes, in order to facilitate the specific binding of liposomes to the receptor of P-selectin expressed on activated platelets. Our results indicate that SuLe(a)-PEG-DSPE can form stable, narrowly distributed liposomes with 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC) and cholesterol, with a vesicle size of 113.3 nm. The resultant SuLe(a)-PEG-liposomes can facilitate their binding to the receptor of P-selectin 22 times higher than SuLe(a)-liposomes without a PEG spacer. Further studies by fluorescence microscopy show that SuLe(a)-PEG-liposomes can bind to activated platelets in vitro effectively. It suggests that biomimetic SuLe(a)-PEG-liposomes may be used as nanocarriers to target activated platelets for drug delivery to the injury sites of cardiovascular diseases.