The microcantilevers have emerged as a versatile biosensor, and showed excellent performance such as high sensitivity, high selectivity, and label-free detection. They have been successfully used for the detection of nucleic acids, disease marker proteins, cells, and pathogens including small molecules. So far, our group has successfully demonstrated the marker protein detection using the actuating layer (PZT)-embedded microcantilevers for the last decade. Here, we introduce in/ex-situ monitoring of the DNA binding events using performance improved actuating layer-embedded microcantilever sensors. To obtain the stable and reliable resonant frequency shifts, the microcantilevers were passivated with parylene-C film for in-situ detection and perfluorosilane (PF-Si) film for ex-situ detection. To achieve the recognition layer, the probe DNA (37-mer including T10 spacers) specific to HBV DNA was immobilized on the gold-coated microcantilever, and followed by backfilling of ethylene glycol spacer (HSC11-EG3-OH) to increase the DNA binding efficiency. After the surface treatment, the detection of HBV DNA (27-mer) was performed through two manners, in-situ and ex-situ. Target DNA in the range of 1 to 20 M and 10 nM to 5 M were applied for the in-situ and ex-situ detection respectively, and the resonant frequency shifts according to the concentration was examined quantitatively. From the results, we explained the relationship between the DNA hybridization and the nanomechanical response. In addition, we presented a hypothesis on the different tendency of in-situ and ex-situ results.