Abstract
We present a nanomechanical platform for real-time quantitative label-free detection of target biomolecules in a liquid environment with mass sensitivity down to few pg. Newly fabricated arrays of up to 18 cantilevers are integrated in a micromachined fluidic chamber, connected to software-controlled fluidic pumps for automated sample injections. We discuss two functionalization approaches to independently sensitize the interface of different cantilevers. A custom piezo-stack actuator and optical readout system enable the measurement of resonance frequencies up to 2MHz. We implement a new measurement strategy based on a phase-locked loop (PLL), built via in-house developed software. The PLL allows us to track, within the same experiment, the evolution of resonance frequency over time of up to four modes for all the cantilevers in the array. With respect to the previous measurement technique, based on standard frequency sweep, the PLL enhances the estimated detection limit of the device by a factor of 7 (down to 2pg in 5min integration time) and the time resolution by more than threefold (below 15s), being on par with commercial gold-standard techniques. The detection limit and noise of the new setup are investigated via Allan deviation and standard deviation analysis, considering different resonance modes and interface chemistries. As a proof-of-concept, we show the immobilization and label-free in situ detection of live bacterial cells (E. coli), demonstrating qualitative and quantitative agreement in the mechanical response of three different resonance modes.
Highlights
In the past 20 years, microcantilever biosensors operated in liquid demonstrated outstanding sensing capabilities.1–4 Several labelfree nanomechanical assays have been developed, targeting the real-time detection of specific biomarkers in physiological environment.5,6 The detection of molecules of clinical interest such as proteins,7 RNA,8–10 and cells11–15 achieved comparable or better performances to commercial gold-standard techniques
We recently demonstrated a direct one-step label-free quantitative immunoassay investigating malaria vaccines, with cantilever arrays with a sensitivity that is on par with the gold-standard multi-step enzyme-linked immunosorbent assay (ELISA) procedure in serum
Microfabricated cantilever arrays present a number of advantages: (i) internal control toward unspecific binding can be achieved by passivating the interfaces of selected sensors; (ii) possibility to perform a differential readout among multiple sensors
Summary
In the past 20 years, microcantilever biosensors operated in liquid demonstrated outstanding sensing capabilities. Several labelfree nanomechanical assays have been developed, targeting the real-time detection of specific biomarkers in physiological environment. The detection of molecules of clinical interest such as proteins, RNA, and cells achieved comparable or better performances to commercial gold-standard techniques. Microfabricated cantilever arrays present a number of advantages: (i) internal control toward unspecific binding can be achieved by passivating the interfaces of selected sensors; (ii) possibility to perform a differential readout among multiple sensors This allows us to correct for thermal drifts or environmental changes, but it makes possible to compare the binding of the same analyte to different surfaces or antigens; (iii) the mechanical response from sensors sensitized with the same biochemical functionalization can be averaged, increasing statistical robustness of experimental results; (iv) possibility to study multiple biochemical interactions within the same experiment; (v) reduction in time and cost of a single test. After Si3N4 and SiO2 removal in wet solutions (hot H3PO4 and 10:1 HF, respectively), shown, the resonator geometry is patterned via a third photolithography followed by silicon dry etching [Fig. 1(f)]. Thinner cantilevers result in a lower spring constant (down to 0.03 N/m), more suitable for static mode operation where quasi-static deflection is targeted
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
