Development of a microfluidic platform for size-based hydrodynamic enrichment and PSMA-targeted immunomagnetic isolation of circulating tumour cells in prostate cancer.

Efforts to further improve the clinical management of prostate cancer (PCa) are hindered by delays in diagnosis of tumours and treatment deficiencies, as well as inaccurate prognoses that lead to unnecessary or inefficient treatments. The quantitative and qualitative analysis of circulating tumour cells (CTCs) may address these issues and could facilitate the selection of effective treatment courses and the discovery of new therapeutic targets. Therefore, there is much interest in isolation of elusive CTCs from blood. We introduce a microfluidic platform composed of a multiorifice flow fractionation (MOFF) filter cascaded to an integrated microfluidic magnetic (IMM) chip. The MOFF filter is primarily employed to enrich immunomagnetically labeled blood samples by size-based hydrodynamic removal of free magnetic beads that must originally be added to samples at disproportionately high concentrations to ensure the efficient immunomagnetic labeling of target cancer cells. The IMM chip is then utilized to capture prostate-specific membrane antigen-immunomagnetically labeled cancer cells from enriched samples. Our preclinical studies showed that the proposed method can selectively capture up to 75% of blood-borne PCa cells at clinically-relevant low concentrations (as low as 5 cells/ml), with the IMM chip showing up to 100% magnetic capture capability.

Smartphone-based colorimetric ELISA implementation for determination of women's reproductive steroid hormone profiles.

Biologists frequently collect and analyze biospecimens in naturalistic (i.e., field) conditions to ascertain information regarding the physiological status of their study participants. Generally, field-collected biospecimens need to be stored frozen in the field and then transported frozen to laboratory facilities where traditional biomarker assays, such as enzyme-linked immunosorbent assays (ELISAs), are conducted. As proper storage and transport of frozen specimens is often logistically difficult and expensive, particularly in nonurban field settings, methods that reduce the need for specimen storage and transport would benefit field-research dependent disciplines such as biology, ecology and epidemiology. One limiting factor to running assays in the field is the use of large and expensive equipment to visualize and quantify the assays, such as microplate readers. Here, we describe an implementation of colorimetric ELISA visualization and quantification using two novel and portable imaging instrumentation systems and data processing techniques for the determination of women's reproductive steroid hormone profiles. Using the light absorbance and transmittance properties of the chemical compounds that make up the hormone assay, we were able to estimate unknown hormone concentrations using a smartphone system and a webcam system. These estimates were comparable to those from a standard laboratory multiple reader (smartphone: accuracy = 82.20%, R 2 > 0.910; webcam: accuracy = 87.59%, R 2 > 0.942). This line of applied research, in the long run, is expected to provide necessary information for examining the extent to which reproductive function varies within and between populations and how it is influenced by psychosocial, energetic and environmental challenges. Our validation of these novel, portable visualization and quantification systems allows for the eventual development of a compact and economical closed system which can be used to quantify biomarker concentrations in remote areas.

An integrated microfluidic chip for immunomagnetic detection and isolation of rare prostate cancer cells from blood.

The quantitative and qualitative analysis of circulating tumor cells (CTCs) has the potential to improve the clinical management of several cancers, including prostate cancer. As such, there is much interest in the isolation of CTCs from the peripheral blood of cancer patients. We report the design, fabrication, and proof-of-principle testing of an integrated permalloy-based microfluidic chip for immunomagnetic isolation of blood-borne prostate cancer cells using an antibody targeting prostate surface membrane antigen (PSMA). The preliminary results using spiked blood samples indicate that the proposed device is consistently capable of isolating prostate cancer cells with high sensitivity (up to 98 %) at clinically relevant low concentrations (down to 20 cells/mL) and an acceptable throughput (100 µL/min).

Detection and isolation of circulating tumor cells: principles and methods.

Efforts to improve the clinical management of several cancers include finding better methods for the quantitative and qualitative analysis of circulating tumor cells (CTCs). However, detection and isolation of CTCs from the blood circulation is not a trivial task given their scarcity and the lack of reliable markers to identify these cells. With a variety of emerging technologies, a thorough review of the exploited principles and techniques as well as the trends observed in the development of these technologies can assist researchers to recognize the potential improvements and alternative approaches. To help better understand the related biological concepts, a simplified framework explaining cancer formation and its spread to other organs as well as how CTCs contribute to this process has been presented first. Then, based on their basic working-principles, the existing methods for detection and isolation of CTCs have been classified and reviewed as nucleic acid-based, physical properties-based and antibody-based methods. The review of literature suggests that antibody-based methods, particularly in conjunction with a microfluidic lab-on-a-chip setting, offer the highest overall performance for detection and isolation of CTCs. Further biological and engineering-related research is required to improve the existing methods. These include finding more specific markers for CTCs as well as enhancing the throughput, sensitivity, and analytic functionality of current devices.

Fungal pathogenic nucleic acid detection achieved with a microfluidic microarray device.

Detection of polymerase chain reaction (PCR) products obtained from cultured greenhouse fungal pathogens, Botrytis cinerea and Didymella bryoniae has been achieved using a previously developed microfluidic microarray assembly (MMA) device. The flexible probe construction and rapid DNA detection resulted from the use of centrifugal pumping in the steps of probe introduction and sample delivery, respectively. The line arrays of the oligonucleotide probes were "printed" on a CD-like glass chip using a polydimethylsiloxane (PDMS) polymer plate with radial microfluidic channels, and the sample hybridizations were conducted within the spiral channels on the second plate. The experimental conditions of probe immobilization and sample hybridization were optimized, and both complementary oligonucleotides and PCR products were tested. We were able to achieve adequate fluorescent signals with a sample load as small as 0.5 nM (1 microL) for oligonucleotide samples; for PCR products, we achieved detection at the level of 3 ng.

An acoustic wave sensor incorporated with a microfluidic chip for analyzing muscle cell contraction.

We report the fabrication of a microfluidic chip or lab-on-a-chip integrated with a thickness-shear mode (TSM) acoustic wave sensor for muscle cell analysis. The sensor, essentially an AT-cut quartz crystal, serves as a detector for recording changes in acoustic wave properties occurring in an attached cardiomyocyte (single heart muscle cell) during its contraction and relaxation. Presumably, the changes resulted from alterations in viscoelastic properties (e.g. stiffness) of the cells. The effects of excitation electrode size, the presence of a microfluidic channel plate, and liquid loading on the sensor were first examined. Thereafter, muscle cell contraction analysis upon chemical stimuli were described. The potential of the chip for screening of cardiovascular drugs is discussed.