Detection of single analyte and environmental samples with silicon nitride nanopores: Antarctic dirt particulates and DNA in artificial seawater

Abstract

Nanopore sensing is a powerful tool for the detection of biomolecules. Solid-state nanopores act as single-molecule sensors that can function in harsh conditions. Their resilient nature makes them attractive candidates for taking this technology into the field to measure environmental samples for life detection in space and water quality monitoring. Here, we discuss the fabrication of silicon nitride pores from ∼1.6 to 20 nm in diameter in 20-nm-thick silicon nitride membranes suspended on glass chips and their performance. We detect pure laboratory samples containing a single analyte including DNA, BSA, microRNA, TAT, and poly-D-lys-hydrobromide. We also measured an environmental (mixed-analyte) sample, containing Antarctic dirt provided by NASA Ames. For DNA measurements, in addition to using KCl and NaCl solutions, we used the artificial (synthetic) seawater, which is a mixture of different salts mimicking the composition of natural seawater. These samples were spiked with double-stranded DNA (dsDNA) fragments at different concentrations to establish the limits of nanopore sensitivity in candidate environment conditions. Nanopore chips were cleaned and reused for successive measurements. A stand-alone, 1-MHz-bandwidth Chimera amplifier was used to determine the DNA concentration in artificial seawater that we can detect in a practical time scale of a few minutes. We also designed and developed a new compact nanopore reader, a portable read-out device with miniaturized fluidic cells, which can obtain translocation data at bandwidths up to 100 kHz. Using this new instrument, we record translocations of 400 bp, 1000 bp, and 15000 bp dsDNA fragments and show discrimination by analysis of current amplitude and event duration histograms.

Protein‐enabled detection of ibuprofen and sulfamethoxazole using solid‐state nanopores

“…Here, we use proteins as carriers and present a method based on SiN x pores for detection of two small drug molecules in the class of pharmaceuticals and personal care products (PPCPs) toward field use (Figure 1C). The SiN x membrane is supported by low-noise (capacitance <1 pF) glass chip [1,14,20,[41][42][43][44]. Pores are reused after rinsing in ethanol [14].…”

“…Nanopore sizes, composition, and properties can be fine-tuned using a range of techniques from electron irradiation [17] to electroporation [18] and down to single-atom-thin pores in a multitude of materials and sub-nm pore diameters [19]. This flexibility makes them amenable to identifying and distinguishing small molecules (e.g., nucleic acids, peptides, and proteins) at low concentrations [1,3,8,14,[20][21][22][23][24][25][26]. As example of environmental applications, nanopores were F I G U R E 1 (A) TEM image 12.5-nm-diameter pore (white circle) in a 20-nm-thick SiN x membrane.…”

“…Scale bar in the image is 2 cm. BSA, bovine serum albumin; SiN x , silicon nitride; SMZ, sulfamethoxazole; TEM, transmission electron microscope used to detect particulates in the Antarctic dirt, DNA in artificial seawater, and biomolecules including proteins, microRNA, and DNA [14].…”

“…BSA have been reported as major binding proteins in blood plasma to different types of small pharmaceutical drugs and are used as the model compound in protein-drug interaction studies (due to its structural homology with human serum albumin and versatile binding activities) [33][34][35][36][37][38]. The ibuprofen and sulfamethoxazole (SMZ) are known to bind the BSA in different sites and are supposed to increase the global negative charge [14,27] of the protein (at higher pH [7][8]. The main hypothesis of this paper is that the molecule binding will affect the translo-…”

The NEOtrap – en route with a new single-molecule technique

“…Different shapes of the nanopore sensor can be used to achieve optimal sensitivity for a given protein effect ( Figure 2 C). For example, longer nanopore channels in thick silicon nitride (SiN) ( Niedzwiecki et al., 2020 ) or polymer membranes ( Buchsbaum et al., 2014 ) provide for a long cavity that is well suited to test, for example, protein fibrillization in a time-resolved way, as further discussed below. The docking position for the origami sphere can also be engineered, for example, with a cup-shaped nanopore opening, to further control the sphere position.…”