Jonathan D. Pegan

Shrinky-Dink microfluidics: rapid generation of deep and rounded patterns

We present a rapid and non-photolithographic approach to microfluidic pattern generation by leveraging the inherent shrinkage properties of biaxially oriented polystyrene thermoplastic sheets. This novel approach yields channels deep enough for mammalian cell assays, with demonstrated heights up to 80 microm. Moreover, we can consistently and easily achieve rounded channels, multi-height channels, and channels as thin as 65 microm in width. Finally, we demonstrate the utility of this simple microfabrication approach by fabricating a functional gradient generator. The whole process--from device design conception to working device--can be completed within minutes.

Tunable shrink-induced honeycomb microwell arrays for uniform embryoid bodies

Embryoid body (EB) formation closely recapitulates early embryonic development with respect to lineage commitment. Because it is greatly affected by cell-cell and cell-substrate interactions, the ability to control the initial number of cells in the aggregates and to provide an appropriate substrate are crucial parameters for uniform EB formation. Here we report of an ultra-rapid fabrication and culture method utilizing a laser-jet printer to generate closely arrayed honeycomb microwells of tunable sizes for the induction of uniform EBs from single cell suspension. By printing various microwell patterns onto pre-stressed polystyrene sheets, and through heat induced shrinking, high aspect micromolds are generated. Notably, we achieve rounded bottom polydimethylsiloxane (PDMS) wells not easily achievable with standard microfabrication methods, but critical to achieve spherical EBs. Furthermore, by simply controlling the size of the microwells and the concentration of the cell suspension we can control the initial size of the cell aggregate, thus influencing lineage commitment. In addition, these microwells are easily adaptable and scalable to most standard well plates and easily integrated into commercial liquid handling systems to provide an inexpensive and easy high throughput compound screening platform.

Flexible shrink-induced high surface area electrodes for electrochemiluminescent sensing

Photolithographically defined metallic thin film on commodity shrink-wrap is leveraged to create robust electrodes. By thermally shrinking the film, electrodes are reduced by 20× in footprint for improved resolution and conductivity with >600% enhancements in electrochemically active surface area; as electrochemiluminescent sensors, they demonstrate improved limits of detection.

Highly stretchable wrinkled gold thin film wires

With the growing prominence of wearable electronic technology, there is a need to improve the mechanical reliability of electronics for more demanding applications. Conductive wires represent a vital component present in all electronics. Unlike traditional planar and rigid electronics, these new wearable electrical components must conform to curvilinear surfaces, stretch with the body, and remain unobtrusive and low profile. In this paper, the piezoresistive response of shrink induced wrinkled gold thin films under strain demonstrates robust conductive performance in excess of 200% strain. Importantly, the wrinkled metallic thin films displayed negligible change in resistance of up to 100% strain. The wrinkled metallic wires exhibited consistent performance after repetitive strain. Importantly, these wrinkled thin films are inexpensive to fabricate and are compatible with roll to roll manufacturing processes. We propose that these wrinkled metal thin film wires are an attractive alternative to conventional wires for wearable applications.

Shrinky-Dink Hanging Drops: A Simple Way to Form and Culture Embryoid Bodies

Embryoid bodies (EB) are aggregates of embryonic stem cells. The most common way of creating these aggregates is the hanging drop method, a laborious approach of pipetting an arbitrary number of cells into well plates. The interactions between the stem cells forced into close proximity of one another promotes the generation of the EBs. Because the media in each of the wells has to be manually exchanged every day, this approach is manually intensive. Moreover, because environmental parameters including cell-cell, cell-soluble factor interactions, pH, and oxygen availability can be functions of EB size, cell populations obtained from traditional hanging drops can vary dramatically even when cultured under identical conditions. Recent studies have indeed shown that the initial number of cells forming the aggregate can have significant effects on stem cell differentiation. We have developed a simple, rapid, and scalable culture method to load pre-defined numbers of cells into microfabricated wells and maintain them for embryoid body development. Finally, these cells are easily accessible for further analysis and experimentation. This method is amenable to any lab and requires no dedicated equipment. We demonstrate this method by creating embryoid bodies using a red fluorescent mouse cell line (129S6B6-F1).

Skin-mountable stretch sensor for wearable health monitoring

This work presents a wrinkled Platinum (wPt) strain sensor with tunable strain sensitivity for applications in wearable health monitoring. These stretchable sensors show a dynamic range of up to 185% strain and gauge factor (GF) of 42. This is believed to be the highest reported GF of any metal thin film strain sensor over a physiologically relevant dynamic range to date. Importantly, sensitivity and dynamic range are tunable to the application by adjusting wPt film thickness. Performance is reliable over 1000 cycles with low hysteresis after sensor conditioning. The possibility of using such a sensor for real-time respiratory monitoring by measuring chest wall displacement and correlating with lung volume is demonstrated.

Superwetting and aptamer functionalized shrink-induced high surface area electrochemical sensors

Electrochemical sensing is moving to the forefront of point-of-care and wearable molecular sensing technologies due to the ability to miniaturize the required equipment, a critical advantage over optical methods in this field. Electrochemical sensors that employ roughness to increase their microscopic surface area offer a strategy to combatting the loss in signal associated with the loss of macroscopic surface area upon miniaturization. A simple, low-cost method of creating such roughness has emerged with the development of shrink-induced high surface area electrodes. Building on this approach, we demonstrate here a greater than 12-fold enhancement in electrochemically active surface area over conventional electrodes of equivalent on-chip footprint areas. This two-fold improvement on previous performance is obtained via the creation of a superwetting surface condition facilitated by a dissolvable polymer coating. As a test bed to illustrate the utility of this approach, we further show that electrochemical aptamer-based sensors exhibit exceptional signal strength (signal-to-noise) and excellent signal gain (relative change in signal upon target binding) when deployed on these shrink electrodes. Indeed, the observed 330% gain we observe for a kanamycin sensor is 2-fold greater than that seen on planar gold electrodes.

Round-bottomed Honeycomb Microwells: Embryoid body shape correlates with stem cell fate

The differentiation of embryonic stem cells (ESC) into tissue-specific cells utilizes either monolayer cultures or three-dimensional cell aggregates called embryoid bodies (EB). However, the generation of a large number of EB of controlled sizes can be challenging and labor intensive. Our laboratories have developed a simple, robust, ultra-rapid, and inexpensive design of Honeycomb Microwells for generation of EB. Here, we compare EB generated using (1) Honeycomb Microwells, (2) the commercially available AggreWell™400, and (3) the more traditional Hanging Drop method. We compared the efficiency, viability, quality, and control of EB sizes. Results indicate that the Honeycomb Microwell and AggreWell™400 efficiently generate small EB at approximately 500 cells per EB. However, the cone-bottomed AggreWell plate generates cone-shaped EB at 1000-2000 cells per EB. Moreover, the cone-shape correlates with a reduction in the formation of the primitive endoderm GATA-4+ cells (1% compared with 6-8% in spherical EB), but does not significantly affect mesoderm or ectoderm development. We conclude that the non-spherical EB shape correlates with a reduction in the development of primitive endoderm, and that use of these AggreWell plates should be avoided in deriving endoderm tissue products. Key words: Embryonic stem cells, embryoid bodies, Microwell, AggreWell, hanging drop, primitive endoderm, GATA-4.

Use of the shape memory polymer polystyrene in the creation of thin film stretchable sensors for wearable applications

The shape memory polymer polystyrene (PS) has been used to create complex hierarchical wrinkling in the fabrication of stretchable thin film bimetallic sensors ideal for wearable based gesture monitoring applications. The film has been bonded to the elastomer polydimethylsiloxane (PDMS) and operates as a strain gauge under the general notion of geometric piezoresistivity. The film was subject to tensile, cyclic, and step loading conditions in order to characterize its dynamic behavior. To measure the joint angle of the metacarpophalangeal (MCP) joint on the right index finger, the sensor was adhered to a fitted golf glove above said joint and a motion study was conducted. At maximum joint angle the sensor experienced roughly 23.5% strain. From the study it was found that two simple curves, one while the finger was in flexion and the other while the finger was in extension, were able to predict the joint angle from measured voltage with an average error of 2.99 degrees.

Smart (shrink manufacturing advanced research tools)

The challenge of micro- and nano-fabrication lies in the difficulties and costs associated with patterning at such high resolution. To make such promising technology - which could enable pervasive health monitoring and disease detection/surveillance - more accessible and pervasive, there is a critical need to develop a manufacturing approach such that prototypes as well as complete manufactured devices cost only pennies. To accomplish this, instead of relying on traditional fabrication techniques largely inherited from the semiconductor industry, we have pioneered a radically different approach. Leveraging the inherent heat-induced relaxation of pre-stressed thermoplastic sheets - commodity shrink-wrap film - we pattern in a variety of ways at the large scale and achieve our desired structures by controlled shrinking down to 5% of the original, patterned sizes.