A microfabricated, high resonance frequency cantilever operating in vacuum with a high quality factor can detect changes in its mass with unprecedented sensitivity into sub-femtogram and beyond. However, severe damping of a resonating cantilever in a liquid media reduces it mass sensitivity when operated in a liquid restricting its use as a mass sensor for biomolecules in their native states. Recent developments in the fabrication of cantilevers, with embedded microfluidic channels, however, enable their operation in vacuum conditions. Since this microfluidic cantilever is operated in vacuum, it allows high sensitivity mass-based detection of biomolecules in the confined liquid inside the fluidic channel. Although biomolecular detection based on mass using microfluidic cantilever is extremely sensitive, it cannot provide any molecular selectivity. Therefore, separation of biomolecules prior to detection is an essential condition for this sensing approach. Separation of biomolecules by capillary gel electrophoresis is well-established method, but it is technically challenging for micron-sized systems such as microfluidic cantilever. For example, uniformly immobilizing the stationary phase inside cantilever channel is a challenge due to its extreme small size. In addition, ultra-low ionic conductivity of integrated channels makes it difficult to realize charge-based separation of mixed molecules even at the time scale of several hours.

However by coupling a microfluidic cantilever with capillary electrophoresis, it is possible to develop a time-of-flow mass spectrometer that can detect separated biomolecules in their native state. Recently we demonstrated such an electrophoresis assisted time-of-flow mass spectrometer using ‘U’ shaped hollow nanomechanical resonators (HNR). By combining an external capillary electrophoresis arrangement with the HNR based mass detection, it is possible to overcome the low ionic conductivity of channels embedded in the HNR preventing direct in-situ electrophoretic separation. The flow of separated biomolecules through the HNR was achieved by balancing the hydrodynamic pressure to override the electromotive force and inhibit the motion of analytes towards the anode for capillary electrophoresis. As the separated biomolecules go through the HNR, the resonance frequency changes sensitively, providing a frequency-time plot (mass-time plot). With a resonance frequency of around 1.5 MHz, the HNR could detect complex samples, such as egg white proteins, in the molecular weight range of 14-250 kDa. Through integrated separation and detection mechanism, this method has the potential to provide precise and fast detection of separated biomolecules in their native state compared to conventional mass spectrometry. This technique also eliminates the need for staining used in conventional polyacrylamide gel electrophoresis. Finally, since the microfluidic cantilevers can be vacuum packaged as a chip, this technique eliminate the need for vacuum pumps, which has been the main bottleneck in the miniaturization of mass spectrometers.

These cantilevers also can be fabricated as bi-material microfluidic cantilevers (BMC) to detect extremely small changes in their temperature. Illuminating the BMC with a tunable wavelength quantum cascade laser (operating in the mid-infrared region) can selectively excite the molecules in the channel. Non-radiative decay of the excited molecules results in the generation of thermal energy, which can be observed as the cantilever bending. A plot of the cantilever bending as a function of the illuminating wavelength shows the IR absorption peaks of the molecules. These nanomechanical IR spectra can be used for unique identification of molecules.

Therefore, by combining the liquid-based micromechanical mass spectrometry with the mid-infrared micromechanical spectroscopy offers a novel platform for molecular recognition of biomolecules in their native state. This concept paves the way towards the development of low-cost and on-chip mass spectrometers with ultra-miniaturized dimensions for field applications where the sample consumption is just under a nanoliter.