Collagen Type IV Alpha 1 (COL4a1) is a key structural component of basement membranes. It forms a network with other type IV collagen chains to provide mechanical stability and organizational scaffolding for tissues. Primarily expressed in vascular, renal, and neural tissues, COL4A1 mutations are associated with a spectrum of disorders, including hereditary angiopathy with nephropathy, aneurysms, and muscle cramps (HANAC) syndrome, as well as porencephaly and hemorrhagic stroke. These mutations disrupt extracellular matrix integrity, leading to vascular fragility and tissue dysfunction. Beyond its structural role, COL4A1 interacts with signaling molecules to regulate cell adhesion and differentiation. Its critical function in tissue development and maintenance makes it a focus of research in genetic and vascular diseases.Besides,Glycoprotein VI (GP6) has been identified as an interactor of COL4a1, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant mouse COL4a1 and recombinant human GP6. Briefly, COL4a1 was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to GP6-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-COL4a1 pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody for 1h at 37℃, wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50 μL stop solution to the wells and read at 450/630nm immediately. The binding activity of recombinant mouse COL4a1 and recombinant human GP6 was shown in Figure 1, the EC50?for this effect is 0.05μg/mL.