HOXB4 protein is a transcription factor encoded by the HOXB4 gene within the HOXB gene cluster of the homeobox gene family.During embryonic development, HOXB4 plays a pivotal role in anterior - posterior (A - P) axis patterning.?In hematopoiesis, HOXB4 is a key regulator. It promotes the self - renewal of hematopoietic stem cells (HSCs). Besides,Sex Determining Region Y Box Protein 2 (SOX2) has been identified as an interactor of HOXB4, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant human HOXB4 and recombinant human SOX2 . Briefly, biotin-linked HOXB4 were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100μl were then transferred to SOX2-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 HOXB4 and SOX2 was shown in Figure 1, the EC50?for this effect is 0.044ug/mL.