multimodal-analysis-microplastics-drinking-water-using-silicon

Multimodal Analysis of Microplastics in Drinking Water using a Silicon Nanomembrane Analysis Pipeline

The biological impact of microplastic pollution in human food and water sources is largely unknown, and drinking water sources are not exempt from this ...

University of Rochester

To reduce unwanted variation in the passage speed of DNA through solid-state nanopores, we demonstrate nanoscale preconfinement of translocating molecules using an ultrathin nanoporous silicon nitride membrane separated from a single sensing nanopore by a nanoscale cavity. We present comprehensive experimental and simulation results demonstrating that the presence of an integrated nanofilter within nanoscale distances of the sensing pore eliminates the dependence of molecular passage time distributions on pore size, revealing a global minimum in the coefficient of variation of the passage time. These results provide experimental verification that the inter- and intramolecular passage time variation depends on the conformational entropy of each molecule prior to translocation. Furthermore, we show that the observed consistently narrower passage time distributions enables a more reliable DNA length separation independent of pore size and stability. We also demonstrate that the composite nanofilter/nanopore devices can be configured to suppress the frequency of folded translocations, ensuring single-file passage of captured DNA molecules. By greatly increasing the rate at which usable data can be collected, these unique attributes will offer significant practical advantages to many solid-state nanopore-based sensing schemes, including sequencing, genomic mapping, and barcoded target detection.

DNA Translocations through Nanopores under Nanoscale Preconfinement.

Silicon nanomembrane technologies (NPN, pnc-Si, and others) have been used commercially as electron microscopy (EM) substrates, and as filters with nanometer-resolution size cut-offs. Combined with EM, these materials provide a platform for catching or suspending nanoscale-size structures for analysis. Usefully, the nanomembrane itself can be manufactured to achieve a variety of nanopore topographies. The size, shapes, and surfaces of nanopores will influence transport, fouling, sieving, and electrical behavior. Electron tomography (ET) techniques used to recreate nanoscale-sized structures would provide an excellent way to capture this variation. Therefore, we modified a sample holder to accept our standardized 5.4 mm × 5.4 mm silicon nanomembrane chips and imaged NPN nanomembranes (50⁻100 nm thick, 10⁻100 nm nanopore diameters) using transmission electron microscopy (TEM). After imaging and ET reconstruction using a series of freely available tools (ImageJ, TomoJ, SEG3D2, Meshlab), we used COMSOL Multiphysics™ to simulate fluid flow inside a reconstructed nanopore. The results show flow profiles with significantly more complexity than a simple cylindrical model would predict, with regions of stagnation inside the nanopores. We expect that such tomographic reconstructions of ultrathin nanopores will be valuable in elucidating the physics that underlie the many applications of silicon nanomembranes.

TEM Tomography of Pores with Application to Computational Nanoscale Flows in Nanoporous Silicon Nitride (NPN).

Selective cellular transmigration across the microvascular endothelium regulates innate and adaptive immune responses, stem cell localization, and cancer cell metastasis. Integration of traditional microporous membranes into microfluidic vascular models permits the rapid assay of transmigration events but suffers from poor reproduction of the cell permeable basement membrane. Current microporous membranes in these systems have large nonporous regions between micropores that inhibit cell communication and nutrient exchange on the basolateral surface reducing their physiological relevance. Here, the use of 100 nm thick continuously nanoporous silicon nitride membranes as a base substrate for lithographic fabrication of 3 µm pores is presented, resulting in a highly porous (≈30%), dual-scale nano- and microporous membrane for use in an improved vascular transmigration model. Ultrathin membranes are patterned using a precision laser writer for cost-effective, rapid micropore design iterations. The optically transparent dual-scale membranes enable complete observation of leukocyte egress across a variety of pore densities. A maximal density of ≈14 micropores per cell is discovered beyond which cell-substrate interactions are compromised giving rise to endothelial cell losses under flow. Addition of a subluminal extracellular matrix rescues cell adhesion, allowing for the creation of shear-primed endothelial barrier models on nearly 30% continuously porous substrates.

Ultrathin Dual-Scale Nano- and Microporous Membranes for Vascular Transmigration Models.

Nanoscale preconfinement of DNA has been shown to reduce the variation of passage times through solid-state nanopores. Preconfinement has been previously achieved by forming a femtoliter-sized cavity capped with a highly porous layer of nanoporous silicon nitride (NPN). This cavity was formed by sealing a NPN nanofilter membrane against a substrate chip using water vapor delamination. Ultimately, this method of fabrication cannot keep a consistent spacing between the filter and solid-state nanopore due to thermal fluctuations and wrinkles in the membrane, nor can it be fabricated on thousands of individual devices reliably. To overcome these issues, we present a method to fabricate the femtoliter cavity monolithically, using a selective XeF etch to hollow out a polysilicon spacer sandwiched between silicon nitride layers. These monolithically fabricated cavities behave identically to their counterparts formed by vapor delamination, exhibiting similar translocation passage time variation reduction and folding suppression of DNA without requiring extensive manual assembly. The ability to form nanocavity sensors with nanometer-scale precision and to reliably manufacture them at scale using batch wafer processing techniques will find numerous applications, including motion control of polymers for single-molecule detection applications, filtering of dirty samples prior to nanopore detection, and simple fabrication of single-molecule nanobioreactors.

Monolithic Fabrication of NPN/SiN Dual Membrane Cavity for Nanopore-based DNA Sensing.

Elucidating the kinetics of DNA passage through a solid-state nanopore is a fertile field of research, and mechanisms for controlling capture, passage, and trapping of biopolymers are likely to find numerous technological applications. Here we present a nanofiltered nanopore device, which forms an entropic cage for DNA following first passage through the nanopore, trapping the translocated DNA and permitting recapture for subsequent reanalysis and investigation of kinetics of passage under confinement. We characterize the trapping properties of this nanodevice by driving individual DNA polymers into the nanoscale gap separating the nanofilter and the pore, forming an entropic cage similar to a "two pores in series" device, leaving polymers to diffuse in the cage for various time lengths, and attempting to recapture the same molecule. We show that the cage results in effectively permanent trapping when the radius of gyration of the target polymer is significantly larger than the radii of the pores in the nanofilter. We also compare translocation dynamics as a function of translocation direction in order to study the effects of confinement on DNA just prior to translocation, providing further insight into the nanopore translocation process. This nanofiltered nanopore device realizes simple fabrication of a femtoliter nanoreactor in which to study fundamental biophysics and biomolecular reactions on the single-molecule level. The device provides an electrically-permeable single-molecule trap with a higher entropic barrier to escape than previous attempts to fabricate similar structures.

Entropic Trapping of DNA with a Nanofiltered Nanopore.

To better understand the origin of microplastics in municipal drinking water, we evaluated 50 mL water samples from different stages of the City of Rochester's drinking water production and transport route, from Hemlock Lake to the University of Rochester. We directly filtered samples using silicon nitride nanomembrane filters with precisely patterned slit-shaped pores, capturing many of the smallest particulates (<20 μm) that could be absorbed by the human body. We employed machine learning algorithms to quantify the shapes and quantity of debris at different stages of the water transport process, while automatically segregating out fibrous structures from particulate. Particulate concentrations ranged from 13 to 720 particles/mL at different stages of the water transport process and fibrous pollution ranged from 0.4 to 8.3 fibers/mL. A subset of the debris (0.2-8.6%) stained positively with Nile red dye which identifies them as hydrophobic polymers. Further spectroscopic analysis also indicated the presence of many non-plastic particulates, including rust, silicates, and calcium scale. While water leaving the Hemlock Lake facility is mostly devoid of debris, transport through many miles of piping results in the entrainment of a significant amount of debris, including plastics, although in-route reservoirs and end-stage filtration serve to reduce these concentrations.

Silicon Nanomembrane Filtration and Imaging for the Evaluation of Microplastic Entrainment along a Municipal Water Delivery Route.

Pollution by microplastics (MPs) is a growing problem that is now well-recognized, as concerning levels of MPs have been found in drinking water, food, and even human tissues. Given the evolving understanding of their toxicological effects on human health, MPs are an area of concern requiring further study. Consequently, there is a need for greater understanding of the performance characteristics of common MP analytical methods and where possible, for standardizing methods and reporting practices. Here, we report our work comparing filtration and imaging properties of five analytical filters suitable for MP capture and analysis. We compared track-etched polycarbonate with (PCTEG) and without gold coating (PCTE), polytetrafluoroethylene (PTFE), porous silicon (PSi), and gold-coated microslit silicon nitride membranes (MSSN-Au). Four of the filter types had a nominal 1.0 μm cut-off, except for PCTEG which had a 0.8 nominal cut-off. We examined the ultrastructure of each membrane type by electron microscopy to understand how their physical properties influence filtration and imaging performance. We compared clean water filtration rates and timed volume passage for each filter in comparison to its porosity and working surface area. We further compared optical microscopy imaging properties for each filter with model MP samples in both bright-field and fluorescent modes with accompanying Nile Red staining. In terms of absolute and surface area-normalized flow rates, our measurements ranked the filters in order of MSSN-Au > PTFE > PCTE > PCTEG > PSi. Similarly, we found MSSN-Au filters compared favorably in terms of optical microscopy performance. Collectively, these data will aid practitioners when choosing analytical filters for MP surveillance and testing.

Article	Year
DNA Translocations through Nanopores under Nanoscale Preconfinement. Nano letters\| PubMed ID: 29087723	2018
TEM Tomography of Pores with Application to Computational Nanoscale Flows in Nanoporous Silicon Nitride (NPN). Membranes\| PubMed ID: 29865242	2018
Ultrathin Dual-Scale Nano- and Microporous Membranes for Vascular Transmigration Models. Small (Weinheim an der Bergstrasse, Germany)\| PubMed ID: 30632319	2019
Monolithic Fabrication of NPN/SiN Dual Membrane Cavity for Nanopore-based DNA Sensing. Advanced materials interfaces\| PubMed ID: 32577337	2019
Entropic Trapping of DNA with a Nanofiltered Nanopore. ACS applied nano materials\| PubMed ID: 32577609	2019
Silicon Nanomembrane Filtration and Imaging for the Evaluation of Microplastic Entrainment along a Municipal Water Delivery Route. Sustainability\| PubMed ID: 36938128	2020
Comparative evaluation of filtration and imaging properties of analytical filters for microplastic capture and analysis. Chemosphere\| PubMed ID: 37127196	2023

Gregory Madejski

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