A kind of aptamer biosensor was prepared to detect adenosine,with magnetic microspheres as the carrier immobilizing adenosine binding aptamer,then assembling adenosine aptamer digoxin labeled part complementary strand of adenosine binding aptamer（report DNA） on the surface.After adding adenosine,the anti-digoxin alkaline phosphatase（ALP）reacted with it,which could result in a change of chemiluminescence（CL） signal.Adenosine was quantified according to the change of the CL intensity.Factors affecting the assembly and recognition process,including amount of magnetic microsphere,aptamer,report DNA and ALP labeled anti digoxin, were studied.The results showed that under the optimal conditions,the concentration of adenosine was linear to CL intensity in the range of 1.0×10-7-1.0×10-3 mol/L with a detection limit of 1.0×10-7 mol/L.The result showed that this method had a better specificity to adenosine than other nucleosides,and was capable of selectively detecting quantities of adenosine in a complex environment（serum）.