This work describes a novel and simple modification of the current microarray format. It reduces the sample/reagent volume to 1 μl and the hybridization time to 500 s. Both 20mer and 80mer oligonucleotide probes and singly labeled 20mer and 80mer targets, representative of the T-cell acute lymphocytic leukemia 1 (TAL1) gene, have been used to elucidate the performance of this hybridization approach. In this format, called shuttle hybridization, a conventional flat glass DNA microarray is integrated with a PMMA microfluidic chip to reduce the sample and reagent consumption to 1/100 of that associated with the conventional format. A serpentine microtrench is designed and fabricated on a PMMA chip using a widely available CO₂ laser scriber. The trench spacing is compatible with the inter-spot distance in standard microarrays. The microtrench chip and microarray chip are easily aligned and assembled manually so that the microarray is integrated with a microfluidic channel. Discrete sample plugs are employed in the microchannel for hybridization. Flowing through the microchannel with alternating depths and widths scrambles continuous sample plug into discrete short plugs. These plugs are shuttled back and forth along the channel, sweeping over microarray probes while re-circulation mixing occurs inside the plugs. Integrating the microarrays into the microfluidic channel reduces the DNA–DNA hybridization time from 18 h to 500 s. Additionally, the enhancement of DNA hybridization reaction by the microfluidic device is investigated by determining the coefficient of variation (CV), the growth rate of the hybridization signal and the ability to discriminate single-base mismatch. Detection limit of 19 amol was obtained for shuttle hybridization. A 1 μl target was used to hybridize with an array that can hold 5000 probes.

In this paper, we report a new method to incorporate 3D scaffold with electrotaxis measurement in the microfluidic device. The electrotactic response of lung cancer cells in the 3D foam scaffolds which resemble the in vivo pulmonary alveoli may give more insight on cellular behaviors in vivo. The 3D scaffold consists of ordered arrays of uniform spherical pores in gelatin. We found that cell morphology in the 3D scaffold was different from that in 2D substrate. Next, we applied a direct current electric field (EF) of 338 mV/mm through the scaffold for the study of cells’ migration within. We measured the migration directedness and speed of different lung cancer cell lines, CL1-0, CL1-5, and A549, and compared with those examined in 2D gelatin-coated and bare substrates. The migration direction is the same for all conditions but there are clear differences in cell morphology, directedness, and migration speed under EF. Our results demonstrate cell migration under EF is different in 2D and 3D environments and possibly due to different cell morphology and/or substrate stiffness.

Real-time observation of cell growth provides essential information for studies such as cell migration and chemotaxis. A conventional cell incubation device is usually too clumsy for these applications. Here we report a transparent microfluidic device that has an integrated heater and a concentration gradient generator. A piece of indium tin oxide (ITO) coated glass was ablated by our newly developed visible laser-induced backside wet etching (LIBWE) so that transparent heater strips were prepared on the glass substrate. A polymethylmethacrylate (PMMA) microfluidic chamber with flow field rectifiers and a reagent effusion hole was fabricated by a ${\text{CO}}_2$ laser and then assembled with the ITO heater so that the chamber temperature can be controlled for cell culturing. A variable chemical gradient was generated inside the chamber by combining the lateral medium flow and the flow from the effusion hole. Successful culturing was performed inside the device. Continuous long-term $\left(>10\text{days}\right)$ observation on cell growth was achieved. In this work the flow field, medium replacement, and chemical gradient in the microchamber are elaborated.

The wound-healing assay is an easy and economical way to quantify cell migration under diverse stimuli. Traditional assays such as scratch assays and barrier assays are widely and commonly used, but neither of them can represent the complicated condition when a wound occurs. It has been suggested that wound-healing is related to electric fields, which were found to regulate wound re-epithelialization. As a wound occurs, the disruption of epithelial barrier short-circuits the trans-epithelial potential and then a lateral endogenous electric field is created. This field has been proved invitro as an important cue for guiding the migration of fibroblasts, macrophages, and keratinocytes, a phenomenon termed electrotaxis or galvanotaxis. In this paper, we report a microfluidic electrical-stimulated wound-healing chip (ESWHC) integrating electric field with a modified barrier assay. This chip was used to study the migration of fibroblasts under different conditions such as serum, electric field, and wound-healing-promoting drugs. We successfully demonstrate the feasibility of ESWHC to effectively and quantitatively study cell migration during wound-healing process, and therefore this chip could be useful in drug discovery and drug safety tests.

We combine a micro-fluidic electric-field cell-culture (MEC) chip with structured-illumination nano-profilometry (SINAP) to quantitatively study the variations of cancer cell filopodia under external direct-current electric field (dcEF) stimulations. Because the lateral resolution of SINAP is better than 150 nm in bright-field image modality, filopodia with diameters smaller than 200 nm can be observed clearly without fluorescent labeling. In the MEC chip, a homogeneous EF is generated inside the culture area that simulates the endogenous EF environment. With this MEC chip–SINAP system, we directly observe and quantify the biased growth of filopodia of lung cancer cells toward the cathode. The epidermal growth factor receptors around the cell edges are also redistributed to the cathodal side. These results suggest that cancer-cell filopodia respond to the changes in EFs in the microenvironment.

This study develops a novel temperature cycling strategy for executing temperature cycling reactions in laser-etched poly(methylmethacrylate) (PMMA) microfluidic chips. The developed microfluidic chip is circular in shape and is clamped in contact with a circular ITO heater chip of an equivalent diameter. Both chips are fabricated using an economic and versatile laser scribing process. Using this arrangement, a self-sustained radial temperature gradient is generated within the microfluidic chip without the need to thermally isolate the different temperature zones. This study demonstrates the temperature cycling capabilities of the reported microfluidic device by a polymerase chain reaction (PCR) process using ribulose 1,5-bisphosphate carboxylase large subunit (rbcL) gene as a template. The temperature ramping rate of the sample inside the microchannel is determined from the spectral change of a thermochromic liquid crystal (TLC) solution pumped into the channel. The present results confirm that a rapid thermal cycling effect is achieved despite the low thermal conductivity of the PMMA substrate. Using IR thermometry, it is found that the radial temperature gradient of the chip is approximately 2 °C mm⁻¹. The simple system presented in this study has considerable potential for miniaturizing complex integrated reactions requiring different cycling parameters.