Cluster Of Differentiation 4 (CD4) is a glycoprotein expressed on the surface of immune cells, primarily T-helper cells, macrophages, and dendritic cells. It serves as a co-receptor for the T-cell receptor (TCR) during antigen presentation, binding to major histocompatibility complex class II (MHC-II) molecules on antigen-presenting cells (APCs). This interaction stabilizes TCR-MHC-II binding and initiates immune signaling. CD4 also acts as a primary receptor for HIV-1, enabling viral entry into host cells.?Besides,Chemokine C-X-C-Motif Receptor 4 (CXCR4) has been identified as an interactor of CD4, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant mouse CD4 and recombinant mouse CXCR4. Briefly, biotin-linked CD4 were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100μl were then transferred to CXCR4-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST 3 times and incubation with Streptavidin-HRP for 30min, then wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50μl stop solution to the wells and read at 450nm immediately. The binding activity of CD4 and CXCR4 was shown in Figure 1, the EC50?for this effect is 2.57μg/mL.