This study demonstrates the miniaturized nanomechanical drug detection of phenytoin using a single free-standing piezoresistive microcantilever biosensor for antiepileptic therapeutic drug monitoring (TDM).Therapeutic drug monitoring is the measurement of specific drugs so as to maintain an appropriate concentration of the medication in the bloodstream[1].It is essential to monitor drug concentration in blood for prevention of side effects when a physician performs clinical management of specific drugs with narrow therapeutic concentration range.A safe therapeutic drug concentration of phenytoin is 10-20 μg/ml for epileptic treatment.Most drugs in TDM have been most analyzed in hospitals by immunoassays such as fluorescence polarization immunoassay (FPIA).However,it requires trained staffs,time-consuming administrative procedures,and fluorescence-based measurement.Being light in weight,free of fluorescence labeling,and highly compatibile with integrated circuits,the microcantilever label-free biosensing technique provides high sensitivity and miniaturization and hence are potentially portable for a point-of-care platform and personal diagnosis.The sensor chip was fabricated by using the MEMS micromachining process.The illustration of the single free-standing piezoresistive microcantilever fabrication process was shown in Figure 1(a).The SEM image of the complete piezoresistive microcantilever was shown in Figure 1(b).Figure 2 demonstrated the full detected responses of the 100 mM self-assembled monolayer biolinker,100 μg/ml immobilized phenytoin antibody,and 100 μg/ml phenytoin drug.The total changes in resistance and surface stress were 0.35 Ω and 2.2 N/m,respectively.As shown in Figure 3,various concentrations of phenytoin drug from 10 to 500 μg/ml were detected in DI water.An increase of change in electrical resistance (ΔR) and surface stresses from corresponded to an increasing concentration of phenytoin drug.The resolution in resistance of 0.005 Ω was obtained.In order to verify the binding specificity of capture antibody and phenytoin drug,100 μg/ml valproic acid of alternative antiepileptic drug was used and no significant signal was observed.This study confirmed that the present microcantilever biosensor achieved high specificity in drug detection.Figure 4 showed the detected results of phenytoin in comparison between the traditional FPIA technique and the present piezoresisitve microcantilever biosensor.The result showed the similar tendency of increasing phenytoin drug concentrations for both techniques in the therapeutic concentrations.In Figure 5,the phenytoin drug detection using the present microcantilever biosensors was successfully demonstrated in a complex body fluid environment of serum that contained proteins,hormones,drugs,antibodies,antigens and others.The complex fluid environment of serum could obstruct the phenytoin drug to bind to the capture antibody on a sensing surface.In summary,electrical drug detection of phenytoin for epileptic treatment was successfully made by using a single free-standing piezoresistive microcantilever biosensor.The miniaturized piezoresistive microcantilever biosensor exhibited its feasibility and realization potential for point-of-care or personal diagnosis in therapeutic drug monitoring.