Detection and Verification of Intestinal and Placental Alkaline Phosphatase Heterodimer in Extracellular Fluid

Due to the lack of relevant detection methods, the existence and function of the intestinal and placental alkaline phosphatase (IAP-PLAP) heterodimer remain largely elusive. Previously, we screened and obtained an aptamer, BG2, which exhibits specific recognition toward the IAP-PLAP heterodimer. Using BG2 as a probe, the heterodimer was found to be highly expressed on the membrane of various tumor cells as well as circulating tumor cells derived from clinical colorectal cancer samples, thus being regarded as a potential tumor marker. However, whether it is shed into extracellular fluid remains unclear. Herein, we developed a BG2-based chemiluminescence assay method with high sensitivity and selectivity for the detection of the IAP-PLAP heterodimer in biological fluids. Furthermore, based on the Na⁺-dependent binding between BG2 and the IAP-PLAP heterodimer, the captured proteins were successfully released and confirmed indeed IAP-PLAP heterodimer, indicating that they can be shed from the cell membrane into the culture medium. It was also found that the concentration of the IAP-PLAP heterodimer in the cell culture medium is closely correlated with its expression level on the cell membrane. Additionally, the levels of the heterodimer both on the cell membrane and in the culture medium were reduced in senescent cells. These results suggest that the IAP-PLAP heterodimer in body fluids may also serve as a disease marker. We further verified that this method can detect the IAP-PLAP heterodimer spiked in plasma samples with good recoveries, thus providing a method for liquid biopsy.