Bio Micro & Nano System Lab

Overview

The Penn State MINIBio Lab aims at developing and applying micro/nano technologies for biological and medical applications. On one hand, we are interested in studying miniaturized devices and systems that can be integrated with biological system in vitro and in vivo. On the other hand, we are eager to apply these technologies for fundamental biological research and clinic diagnosis and treatment. The research is highly multidisciplinary, interfacing at engineering, biological sciences, physical sciences, and medicine. Current research focuses of the group include:

Micro/nano materials, devices and systems for liquid biopsy-based cancer diagnosis, including enrichment and analysis of circulating tumor cells (CTCs) and extracelluar vesicles (EVs)
Nanomaterials and nanomaterial integrated microdevices for infectious disease diagnosis and pathogen discovery
Nanomaterials and implantable devices for drug delivery and disease monitoring
Nanomaterials for proteomics including peptide capture and ultrafast protein digestion
Lab-on-chip biomarker detection in body fluid
High-performance devices for gas chromatography (GC) and volatile organic compound (VOC) detection

Examples of previous research projects are listed as below:

Cancer Diagnosis by Liqid Biopsy

Project: EV isolation and analysis by lipid nanoprobes

EVs can mediate intercellular communication by transferring cargo proteins and nucleic acids between cells. The pathophysiological roles and clinical value of EVs are under intense investigation, yet most studies are limited by technical challenges in the isolation of EVs. We developed a nanomaterial-based method for EV isolation. The lipid nanoprobe enables spontaneous labelling and magnetic enrichment of EVs in 15 minutes, with high isolation efficiency and purity. We also show that the lipid nanoprobes, which allow for downstream analyses of nucleic acids and proteins, enabled the identification of DNA mutations following EV isolation from blood plasma from cancer patients. The efficiency and versatility of the lipid nanoprobe opens up opportunities in point-of-care cancer diagnostics.

References:

Rapid isolation of extracellular vesicles via lipid nanoprobes, Y. Wan, G. Cheng, X. Liu, S.-J. Hao, M. Nisic, C.-D. Zhu, Y.-Q.Xia, W.-Q. Li, Z.-G. Wang, W.-L. Zhang, S. J. Rice, A. Sebastian, I. Albert, C. P. Belani, S.-Y. Zheng, Nature Biomedical Engineering, 1, 0058, 2017. (Feature as front cover story) [Link] [Behind the Paper] [PSU News Release]

Project: Flexible micro spring array (FMSA) for viable circulating tumor cell (CTC) enrichment

Circulating Tumor Cell enrichment and analysis

We demonstrated a high throughput versatile platform capable of isolating circulating tumor cells (CTCs) from clinically relevant volumes of blood while preserving their viability and ability to proliferate. The enrichment is based on the fact that CTCs are larger compared with normal blood cells. The incorporated system allows size-based separation of CTCs at the micro-scale, while taking advantage of a high throughput and rapid processing speed. Testing results of model systems using cell lines show that this device can enrich CTCs from 7.5 mL of whole blood samples with 90% capture efficiency, higher than 10⁴ enrichment, and better than 80% viability in approximately ten minutes without any incidence of clogging.

References:

Size-based separation methods of circulating tumor cells, S.-J. Hao, Y. Wan, Y.-Q. Xia, X. Zou, and S.-Y. Zheng, Advanced Drug Delivery Reviews, 25, 3-20, 2018. [Link]
Evaluating a novel dimensional reduction approach for mechanical fractionation of cells using a tandem flexible micro spring array (tFMSA), Y. T. Yeh, R. A. Harouaka, S. -Y. Zheng, Lab on a Chip, 17 (4), 691-701, 2017. [Link]
Flexible micro spring array device for high throughput enrichment of viable circulating tumor cells, R. A. Harouaka, M.-D. Zhou, Y.-T. Yeh, W. J. Khan, A. Das, X. Liu, C. C. Christ, D. T. Dicker, T. S. Baney, J. T. Kaifi, C. P. Belani, C. I. Truica, W. S. El-Deiry, J. P. Allerton, and S.-Y. Zheng, Clinical Chemistry, vol. 60, pp. 323-333, 2014. [pdf]
Separable bilayer microfiltration device for viable label-free enrichment of circulating tumour cells, M.-D. Zhou†, S. Hao†, A. J. Williams†, R. A. Harouaka, B. Schrand, S. Rawal, Z. Ao, R. Brennaman, E. Gilboa, B. Lu, S. Wang, J. Zhu, R. Datar, R. Cote, Y.-C. Tai*, and S.-Y. Zheng*, Scientific Reports, 4: 7392, 1-10, doi:10.1038/srep07392, 2014. (†Authors with equal contribution) [pdf]
3D microfilter device for viable circulating tumor cell (CTC) enrichment from blood, S. Zheng, H. K. Lin, B. Lu, A. Williams, R. H. Datar, R. J. Cote RJ, Y. –C. Tai, Biomedical Microdevices, vol.13, pp. 203-213, 2011. [Link]
Portable filter-based microdevice for detection and characterization of circulating tumor cells, H. K. Lin*, S. Zheng*, A. J. Williams, M. Balic, S. Groshen, H. I. Scher, M. Fleisher, W. Stadler, R. H. Datar, Y.-C. Tai, R. J. Cote, Clinical Cancer Research, vol 16, pp. 5011-5018, 2010. (*Authors with equal contribution)
Membrane microfilter device for selective capture, electrolysis and genomic analysis of human circulating tumor cells, S. Zheng*, H. Lin*, J.-Q. Liu, M. Balic, R. Datar, R. J. Cote, and Y.-C. Tai, Journal of Chromatography A, vol. 1162, pp. 154-161, 2007. (*Authors with equal contribution)

Virus discovery and detection

Viral infectious diseases can cause unpredictable but recurring outbreaks that lead to devastating mortality and traumatic economic loss. Almost all lethal viral outbreaks in the past two decades were caused by newly emerging viruses and often zoonotic. Many samples originate from remote areas with limited resources and poor logistic networks, therefore high performance point-of-care device and system for virus sample preparation and detection is highly valuable.

Project: Nanomaterial-integrated microdevice for size-based virus capture

Most of the discovered human viral pathogens have sizes ranging from 20 nm to 300 nm. This is a unique size range that between micrometer-sized bacteria and mammalian cells, and biological small molecules and macromolecules (e.g. proteins, nucleic acids). Recently we invented nanomaterial-integrated microdevice technologies that enriches virus particles unbiasly from field and clinical samples.
In one case (top), we integrated vertically aligned carbon nanotube (VACNT) nanostructures silicon substrate to achieve label-free virus capture. We can control the inter-tubular distance between CNTs in the range between 25 nm to over 300 nm, perfectly matching the size range of viruses. The unique structure of the VACNT forest presents high porosity of over 90%, which minimizes flow resistance, increases sample capacity, and minimizes device clogging. The technology can prepare field and clinical samples for virus discovery. Using the microdevice, we successfully identified an emerging avian influenza virus (AIV) from swab samples and a new strain of infectious bursal disease virus (IBDV) from a small piece of turkey tissue.
In another example (bottom), we developed a microfluidic device integrated with biodegradable silicon nanowires for virus capture and analysis. The device enables operation in continuous flow mode and virtually eliminates channel clogging. In addition, viruses can be released later and collected at the outlet by degrading the porous silicon nanowires in phosphate buffered saline for 24 hours. With this system, we demonstrated label-free AIV capture and release.

References:

Tunable and label-free virus enrichment for ultrasensitive virus detection using carbon nanotube arrays, Y.-T. Yeh, Y. Tang, A. Sebastian, A. Dasgupta, N. Perea-Lopez, I. Albert, H. Lu, M. Terrones, and S.-Y. Zheng, Science Advances, 2, e1601026, 2016. [Link] [pdf] [video] [PSU News Release] [NIH Director's blog]
Label-free virus capture and release by a microfluidic device integrated with porous silicon nanowire forest, Y. Xia, Y. Tang, X. Yu, Y. Wan, Y. Chen, H. Lu, S.-Y. Zheng, Small, 13 (6), 1603135, 2017. (Feature as inside front cover story) [Link]

Project: Nanomaterial-integrated microdevice for virus detection

For targeted virus diagnosis, nanomaterial-integrated microdevice can boost detection sensitivity and selectivity. We developed a nanomaterial-integrated microdevice with surface conjugated antibodies to capture specific viruses and detect them by on-chip ELISA assay. 3D randomly oriented zinc oxide (ZnO) nanorods were fabricaed into several branched microchannels on glass surface for multiplex virus detection. The 3D nanostructure not only provides more binding sites for the virus capture, but also enhances fluorescence detection. The captured viruses can be released by simply dissolving the ZnO nanorods at slightly acidic environment.

References:

A Nanostructured Microfluidic Immunoassay Platform for Highly Sensitive Infectious Pathogen Detection, X. Yu, Y. Xia, Y. Tang, W.-L. Zhang, Y.-T. Yeh, H. Lu & S.-Y. Zheng, Small, 13, 1700425, 2017. (Feature as inside front cover story) [Link] [Nanowerk]
Highly sensitive DNA detection using cascade amplification strategy based on hybridization chain reaction and enzyme-induced metallization, X. Yu, Z.-L. Zhang and S.-Y. Zheng, Biosens. Bioelectron., 66, 520-526, 2015. [Link]

Nanomedicine

Project: EV-camouflaged nanoparticles for protein delivery

The intracellular delivery of biofunctional enzymes or therapeutic proteins through systemic administration is of great importance in therapeutic intervention of various diseases. However, current strategies face substantial challenges owing to various biological barriers, including susceptibility to protein degradation and denaturation, poor cellular uptake, and low transduction efficiency into the cytosol. We developed a biomimetic nanoparticle platform for systemic and intracellular delivery of proteins. Through a biocompatible strategy, guest proteins are caged in the matrix of metal-organic frameworks (MOF) with high efficiency and high loading capacity. The nanoparticles were further enveloped with the extracellular vesicle (EV) membrane. In vitro and in vivo study manifests that the EV-like nanoparticles can not only protect proteins against protease digestion and evade the immune system clearance but also selectively target homotypic tumor sites, promote tumor cell uptake and autonomous release of the guest protein after internalization. Assisted by biomimetic nanoparticles, intracellular delivery of bioactive therapeutic protein gelonin significantly inhibits the tumor growth in vivo.

References:

Self-assembly of extracellular vesicle-like metal–organic framework nanoparticles for protection and intracellular delivery of biofunctional proteins, G. Cheng, W. Li, L. Ha, X. Han, S. Hao, Y. Wan, Z. Wang, F. Dong, X. Zou, Y. Mao, & S.-Y. Zheng, Journal of the American Chemical Society, 140, 7282-7291, 2018. (Feature as supplementary cover story) [Link] [PSU News Release]

Project: Aptamer conjugated EVs for targeted drug delivery

The extracellular vesicles (EVs) have emerged as important disease biomarkers and therapeutic targets, but also have been utilized as novel drug delivery vehicles as EVs are dowered with small size, favorable surface protein composition, and membrane of the parental cells. However, clinical translations of EVs in targeted therapeutics are limited by their low yield and inadequate targeting effect. To produce large amount of EVs, cells grafted with lipidated targeting ligand (aptmer) were mechanically extruded to generate cancer cell targeting artificial EVs in ~1 hour. This rapid and economic approach could pave the way for clinical EV implementation in the future.

References:

Aptamer conjugated extracellular nanovesicles for in vivo targeted drug delivery, Y. Wan, L. Wang, C. Zhu, Q. Zheng, G. Wang, J. Tong, Y. Fang, Y.-Q. Xia, G. Cheng, X. He, and S.-Y. Zheng, Cancer Research, 78, 798-808, 2018. [Link]

Project: Cellular interactions

Nanomedicine-based drug delivery systems can have designed and sometimes unexpected interactions with cells. We studied using dopamine-derived polydopamine nanoparticles with a mitochondrial penetration molecule for mitochondria-targeted drug delivery to overcome drug resistance in cancer therapy (Figure a). In another case (Figure b), we discovered the preoccupation of endo-/lysosomes by the empty nanocarriers has a "saturation" effect mainly through changing the spatial distribution of the subsequently introduced nanocarriers. In the 3rd example (Figure c), we explored using mitochondria as a natural delivery system of carbon quantum dots for in vivo imaging and administration of the anticancer drug doxorubicin.

References:

Mitochondria-Targeting Polydopamine Nanoparticles to Deliver Doxorubicin for Overcoming Drug Resistance, W. Li, Z. Wang, S.-J. Hao, H.He, Y. Wan,C. Zhu, L. Sun, G. Cheng & S. -Y. Zheng, ACS Applied Materials & Interfaces, 9, 16793–16802, 2017.) [Link]
Pre-occupation of empty carriers decreases endo/lysosome escape and reduces the protein delivery efficiency of mesoporous silica nanoparticles, W. Li, L. Sun, Y. Xia, S.-J. Hao, G. Cheng, Z. Wang, Y. Wan, C. Zhu, H. He, and S.-Y. Zheng, ACS Applied Materials & Interfaces, 10, 5340-5347, 2018. [Link]
Mitochondria-based aircraft carrier enhances in vivo imaging of carbon quantum dots and delivery of anticancer drug, W.-Q. Li, Z. Wang, S. Hao, L. Sun, M. Nisic, G. Cheng, C. Zhu, Y. Wan, L. Ha, L. & S.-Y. Zheng, Nanoscale, 10, 3744-3752, 2018. [Link]

Nanomaterials for Proteomics

Project: Nanomaterials for peptide capture

We designed and constructed a novel graphene composite affinity material consisting of graphene scaffolds, Fe₃O₄ nanoparticles for actuation and fully covered porous titania nanostructures as affinity coating. The multifunctional graphene composites can realize the selective capture and convenient magnetic separation of target phosphopeptides by taking advantage of the decorated magnetic nanoparticles, highly pure and well crystallized affinity coating, and unique porous structure. The affinity graphene composites can realize selective capture and rapid separation of low-abundant phosphopeptides from complex biological samples.
In another study, we prepared mesoporous and hollow carbon microspheres embedded with magnetic nanoparticles via a facile self-sacrificial method for rapid capture of low-abundant peptides from complex biological samples. The effectiveness of these affinity microspheres for capture of low-concentration peptides was evaluated by standard peptides, complex protein digests, and real biological samples. These multifunctional hollow carbon microspheres
can realize rapid capture and convenient separation of low-concentration peptides. They were validated to have better performance than magnetic mesoporous silica and commercial peptide-enrichment products. In addition, they can be easily recycled and present excellent reusability.

References:

Facile synthesis of magnetic mesoporous hollow carbon microspheres for rapid capture of low-concentration peptides, G. Cheng, M.-D. Zhou, S.-Y. Zheng, ACS Applied Materials & Interfaces, 6, 12719-12728, 2014. [Link]
Preparation of magnetic graphene composites with hierarchical structure for selective capture of phosphopeptides, G. Cheng, X. Yu, M. Zhou, and S.-Y. Zheng, J. Mater. Chem. B, 2, 4711-4719, 2014. [Link]
Graphene-Templated Synthesis of Magnetic Metal Organic Framework Nanocomposites for Selective Enrichment of Biomolecules, G. Cheng, Z. G. Wang, S. Denagamage, and S.-Y. Zheng, ACS Appl Mater Interfaces, 8, 10234-10242, 2016. [Link]

Project: Nanomaterials for ultrafast protein digestion

We designed a magnetic enzyme nanosystem have by a polydopamine (PDA)-modification strategy (Figure a). The magnetic enzyme nanosystem has well defined core-shell structure and a relatively high saturation magnetization. The magnetic enzyme system can realize rapid, efficient and reusable tryptic digestion of proteins by taking advantage of its magnetic core and biofunctional shell. The magnetic enzyme nanosystem can digest the proteins in 30 minutes, and the results are comparable to conventional 12 hours in-solution digestion. The material can be reused several times, and has excellent stability for storage.
We further implement a similar strategy within a novel microfluidic protein digestion system with nanostructured and bioactive inner surface by an easy biomimetic covalent self-assembly strategy (Figure b). The microfluidic digestion system can realize rapid and effective proteolysis in 2 minutes, which is dramatically faster than the conventional overnight digestion methods by at least two orders of magnitude. Because of this advancement and the advantages of microfluidic chips, it is expected that this work would contribute to rapid online digestion in future high-throughput proteomics. Remote and noninvasive modulation of protein activity is essential for applications in biotechnology and medicine. In the 3rd example (Figure c), a near-infrared (NIR) light responsive graphene-silica-trypsin nanoreactor is developed for modulating the bioactivity of trypsin molecules. Biomolecules are spatially confined and protected in the rationally designed compartment architecture, which not only reduces the possible interference but also boosts the bioreaction efficiency. Upon NIR irradiation, the photothermal effect of the nanoreactor enables the ultrafast in situ heating for remote activation and tuning of the bioactivity. We apply the GST nanoreactor for remote and ultrafast proteolysis of proteins, which remarkably enhance the proteolysis efficiency and reduces the bioreaction time from the overnight of using free trypsin to seconds.

References:

Construction of a high-performance magnetic enzyme nanosystem for rapid tryptic digestion, G. Cheng and S.-Y. Zheng, Scientific Reports, 4: 6947, 1-10, doi:10.1038/srep06947, 2014. [pdf]
Nanostructured microfluidic digestion system for rapid high-performance proteolysis, G. Cheng, S.-J. Hao, X. Yu and S.-Y. Zheng, Lab on Chip, 15, 650-654, doi:10.1039/C4LC01165A, 2015. [Link]
In situ caging of biomolecules in graphene hybrids for light modulated bioactivity, G. Cheng, X.-H. Han, S.-J. Hao, M. Nisic, and S.-Y. Zheng, ACS Applied Materials & Interfaces, 10, 3361-3371, 2018. [Link]

Implantable Microdevices

Project: Implantable pressure sensor for Left Ventricular Assist Device (LVAD)

Image of implantable pressure sensor for Left Ventricular Assist device

Continuous flow left ventricular assist devices (LVADs) are commonly used as bridge-to-transplantation or destination therapy for heart failure patients. However, non-optimal pumping speeds can reduce the efficacy of circulatory support or cause dangerous ventricular arrhythmias. Optimal flow control for continuous flow LVADs has not been defined and calls for an implantable pressure sensor integrated with the LVAD for real-time feedback control of pump speed based on ventricular pressure. A MEMS pressure sensor prototype is designed, fabricated and seamlessly integrated with LVAD to enable real-time control, optimize its performance and reduce its risks. The pressure sensing mechanism is based on Fabry-Pérot interferometer principle. A biocompatible parylene diaphragm with a silicon mirror at the center is fabricated directly on the inlet shell of the LVAD to sense pressure changes. The sensitivity, range and response time of the pressure sensor are measured and validated to meet the requirements of LVAD pressure sensing.

References:

An Implantable Fabry–Pérot Pressure Sensor Fabricated on Left Ventricular Assist Device for Heart Failure, M. D. Zhou, C. Yang, Z. Liu, J. P. Cysyk, and S. Y. Zheng, Biomedical Microdevices, vol.14, pp. 235-245, 2012. [Link]

Blood Analysis

Project: Point-of-care device and system for blood coagulation measurement

Blood coagulation tests are an integral part of diagnosis and treatment of patients with cardiac disorders. The activated partial thromboplastin time test (aPTT or APTT) is performed on a large scale, in laboratories, to monitor the functioning of the intrinsic and common pathways of the blood coagulation cascade and the concentration of anticoagulants such as heparin among patients. A polydimethylsiloxane (PDMS) based microfluidic device has been fabricated and tested to perform APTT as a point-of-care device using whole blood samples by detecting the change in electrical impedance of blood during coagulation. A portable setup consisting of the device and a lock-in amplifier circuit was built and tested for repeatability and stand alone usage. A frequency sweep test was performed on the device and the optimal frequency of operation for maximum output sensitivity was determined. The devices were also tested for their sensitivity to different concentrations of heparin and found to exhibit an expected increase in coagulation time for increasing heparin concentration.

Reference:

Microfluidic device and system for point-of-care blood coagulation measurement based on electrical impedance sensing, B. Ramaswamy, Y. T. Yeh, and S. Y. Zheng, Sensors and Actuators B: Chemical, vol. 14(1), pp. 235-245, 2012. [Link]

MEMS and microfluidic devices

Project: High temperature MEMS valves for GC-based VOC detection

We designed and fabricated a MEMS flow control device for gas chromatography (GC) with the capability of sustaining high-temperature environments. We demonstrated the use of this new device in a novel MEMS chopper-modulated gas chromatography-electroantennography (MEMS-GC-EAG) system to identify specific volatile organic compounds (VOCs) secreted by insects at extremely low concentrations.

Reference:

Chopper-Modulated gas chromatography electroantennography enabled using high-temperature MEMS flow control device, M.-D. Zhou, M. Akbar, A. J. Myrick, Y. Xia, W. J. Khan, X. Gao, T. C. Baker and S.-Y. Zheng, Microsystems & Nanoengineerin, Nature Publishing Group, 3, 17062, 2017. [Link] [pdf]

Project: Microfluidic device for functional microsphere synthesis

We developed a simple and robust method for one-step synthesis of monodisperse functional polymeric microspheres by generation of reversed microemulsion droplets in aqueous phase inside a microfluidic device and controlled evaporation of the organic solvent. Using this method, water-soluble nanomaterials can be easily encapsulated into biodegradable Poly(D, L-lactic-co-glycolic acid) (PLGA) to form functional microspheres. As a demonstration of the versatility of the approach, high-quality fluorescent quantum dots of various emission spectrums, superparamagnetic iron oxide nanoparticles and water-soluble carbon nanotubes were used to synthesize fluorescent, magnetic and CNT-containing polymeric microspheres, respectively.

Reference:

On-Demand One-Step Synthesis of Monodisperse Functional Polymeric Microspheres with Droplet Microfluidics, X. Yu, G. Cheng, M.-D. Zhou & S.-Y. Zheng, Langmuir, 31, 3982-3992, 2015. [Link]

Project: Bone chip to study bone metastasis

Bone metastasis occurs at approximately 70% frequency in metastatic breast cancer and most women who die of metastatic breast cancer have bone metastases. We developed a bone-on-a-chip for spontaneous growth of a 3D, mineralized, collagenous bone tissue. Based on the principle of simultaneous-growth-dialysis, mature osteoblastic tissue containing heavily mineralized collagen fibers naturally formed in 30 days without the aid of differentiation agents. Moreover, we examined co-culture of metastatic human breast cancer cells with osteoblastic tissues. The new bone-on-a-chip design not only increases experimental throughput while reducing fabrication cost by miniaturization, but also maximizes the chances of cancer cell interaction with bone matrix of a concentrated surface area and facilitates easy, frequent observation.

Reference:

A spontaneous 3D bone-on-a-chip for bone metastasis study of breast cancer cells, S. Hao, L. Ha, G. Cheng, Y. Wan, Y. Xia, D. M. Sosnoski, A. M. Mastro, and S.-Y. Zheng, Small, 14, 1702787, 2018. [Link] [Physicsworld]