Our recent study revealed that fibrin interacts with the very low density lipoprotein receptor (VLDLR) on endothelial cells through its βN domains, and this interaction promotes transendothelial migration of leukocytes and thereby inflammation. The major aims of this study were to further characterize this interaction and localize the fibrin-binding site in the VLDLR. To localize the fibrin-binding site, we expressed a soluble extracellular portion of this receptor, sVLDLRHT, its N- and C-terminal regions, VLDLR(1-8)HT and des(1-8)VLDLRHT, respectively, and a number of VLDLR fragments containing various combinations of CR domains and confirmed their proper folding by fluorescence spectroscopy. Interaction of these fragments with the (β15-66)2 fragment corresponding to a pair of VLDLR-binding βN domains of fibrin was tested by different methods. Our experiments performed by an enzyme-linked immunosorbent assay and surface plasmon resonance revealed that the VLDLR(1-8)HT fragment containing eight CR domains of VLDLR and its subfragments, VLDLR(1-4)HT and VLDLR(2-4)HT, interact with (β15-66)2 with practically the same affinity as sVLDLRHT while the affinity of VLDLR(2-3)HT was ∼2-fold lower. In contrast, des(1-8)VLDLRHT exhibited no binding. Formation of the complex in solution between the fibrin-binding fragments of VLDLR and (β15-66)2 was detected by fluorescence spectroscopy. In addition, formation of a complex between VLDLR(2-4)HT and (β15-66)2 in solution was confirmed by size-exclusion chromatography. Thus, the results obtained indicate that minimal fibrin-binding structures are located within the second and third CR domains of the VLDL receptor and the presence of the fourth CR domain is required for high-affinity binding. They also indicate that tryptophan residues of CR domains are involved in this binding.