Multimodal real-time frequency tracking of cantilever arrays in liquid environment for biodetection: Comprehensive setup and performance analysis

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 2 MHz. 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 2 pg in 5 min integration time) and the time resolution by more than threefold (below 15 s), 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.

Quantitative epitope analysis reveals drastic 63% reduced immuno-affinity and 60% enhanced transmissibility for SARS-CoV-2 variants

“… 22 In order to ensure temperature stability and insulation, the whole setup (including fluidic pumps, pipes and valves) is enclosed in a thermally insulated box (stable internal temperature with 0.02 °C precision). 27 Dynamic mode operation is enabled via a custom-built piezo-ceramic stack actuator, placed in a pocket underneath the chip, isolated from the fluidic volume by a 200 μm-thick PEEK membrane. 22 Mechanical signals are detected via optical beam deflection readout.…”

Rapid Label-free Nanotechnological Immunoassay for Analysis of Candidate Malaria Vaccines

“…The minimum detectable mass is given if one takes the minimum measurable frequency change. The sensors are exposed synchronously to the same sample and multiples of individual functionalizations [15] allow for averaging of the specific mechanical response. A minimum of two mechanically equivalent sensors is required [16], we currently use 18 in parallel [15].…”

“…The sensors are exposed synchronously to the same sample and multiples of individual functionalizations [15] allow for averaging of the specific mechanical response. A minimum of two mechanically equivalent sensors is required [16], we currently use 18 in parallel [15]. Synchronised differential readout with in-situ reference probes is mandatory to avoid convolution with external environmental factors.…”

“…Synchronised differential readout with in-situ reference probes is mandatory to avoid convolution with external environmental factors. Such differential analysis also eliminates thermal experimental drifts, flow-induced disturbances, and unspecific interactions as present in serum samples (see Figure 1) [1,13,[15][16][17][18][19][20][21][22][23].…”

“…The epitope-specific antibody recognition (panel a) (red antibodies towards UK39 and grey antibodies as unspecific interactions) generates a frequency shift based on the different sensor functionalizations (panel b). Taking the characteristics of the fluid around the sensors and the sensors' dimensions into account, the frequency shifts are converted into a mass addition towards one or the other epitope (panel c)[15,17,20,22,23].…”