Year: 2020

Impressive research coming out of the Revzin group at the Mayo Clinic’s Department of Physiology and Biomedical Engineering shows an improved approach to first culture and then evaluate cell responses to chemotherapy treatments using polydimethyl siloxane (PDMS) microfluidic devices cast from silicon masters.  The research was published in Microsystems & Nanoengineering in October (DOI: 10.1038/s41378-020-00201-6).

Ovarian cancer often has a very poor prognosis when detected due in large part to the inaccessibility of the ovaries for many diagnostic methods, and the late-stage in which diagnoses are usually made.  As a result, there is a strong focus on improved therapeutic strategies to best use the brief window available for treatment of the advanced disease.  One such strategy is to use ‘patient-derived xenografts’ (PDXs) where tumour tissue cells are implanted in a mouse to allow the cancer to be monitored in parallel to that of the patient.  While valuable to determine the effectiveness of various chemotherapies, PDX requires research animals, experienced animal researchers, high costs and long timelines.  As a result, in vitro ‘organoid’ or ‘spheroid’ 3-D cancer cultures are being explored on 96-well microtiter plates, Matrigel plates and other array platforms to arrive at suitable replacement environments.  As comparison standards for their microfluidic design, the authors used Matrigel and microtiter plates approaches, with the latter having PDMS inserts with microwells identical in size to the microfluidic counterpart.  A graphic depicting the three approaches is shown above at right.

A first advantage of their microfluidic platform is that it can easily make use of the miniscule amounts of cells afforded by a fine needle biopsy sample.  Secondly, the on-chip incubation environment appears to be superior to that of standard microtiter plates when using identical media and culturing methods.  After 14 days, 0n-chip viability was typically ~90% vs. ~60% & ~80% for standard microtiter and Matrigel plates, respectively; growth of the spheroid cluster of cancer cells was also significantly better using the chip platform. Figure at right shows micrographs of the spheroid cell clusters in the microfluidic wells (a), and a comparison of viability (b) and spheroid size growth multiple (c) over 2 weeks.  Lastly, a multiplexed design shown below at right was created to allow serial and parallel perfusion across eight chambers.  Chambers are filled serially from left as in (a), and in parallel via discrete inputs as in (b); fluidic control is achieved by opening and closing red valves surrounding each chamber.  This device was used to determine the chemotherapy agent doxorubicin’s IC50 (50% inhibitory concentration) for two types of cancer cells via viability curves drawn from parallel injections of agent concentrations ranging from 10 nM to 100 µM.  The injected amount for the multiplexed study was ~100,000 cells, stated as the minimum amount obtained from a fine needle aspirate (biopsy).

Application of this microfluidic analysis approach to very small masses of biopsy, or other, samples is compelling as demonstrated in the case of ovarian cancer.  It may be that many disease biopsies from other organs or perhaps forensic or other bio-samples could also benefit from the efficient approach.

A very thorough characterisation paper was published a few weeks ago in Analytical Chemistry by researchers in Amal Alachkar’s group at UC Irvine, and Nicolas Voelcker‘s and Victor Cadarso‘s groups at Monash University.  The collaboration made a microfluidic dopamine sensor that detected dopamine in blood or cerebrospinal fluid (CSF) obtained from mice used to model Parkinson’s disease.  Dopamine is a small molecule that acts as a neurotransmitter in the brain and as a hormone with different functions elsewhere in the body including blood vessels, kidneys, the pancreas and the immune system.

The microfluidic devices were fairly simple in structure, with a three-electrode configuration used for the amperometric detection patterned on the oxide layer of a silicon substrate, and a single channel/chamber fluidic structure fabricated in PDMS that was bonded to the silicon baseplate.

Characterisation tests showed robust performance.  Good repeatability was seen across ten replicate devices, linear ranges extended through 4 decades and included an LoD at 0.1 nM in both CSF and phosphate buffer.  The current plots showing the dynamic range appeared quite steady and smooth at 0.1 nM, and well apart from the baseline, so I wonder if a 3-standard deviation mark would lie well below 0.1 nM?  Regardless, this was adequate to easily sense the physiological levels of dopamine in the test mice CSF in the range of 0.1-1 nM.  It also showed good selectivity in the presence of interferents.  The good analytical performance is likely attributable to the small 2.4 µL chamber, relatively large electrode array (~1 mm²) and narrow (50 µm) separation between the electrodes.

The ability to detect dopamine is of great interest, since abnormal dopamine levels are indicative of neurological conditions such as depression, schizophrenia, attention deficit hyperactivity disorder (ADHD) and Parkinson’s disease.  In the case of Parkinson’s disease, an effective sensor can help both with diagnosis and with treatment, as patients are often treated with  L-DOPA, a chemical precursor to dopamine, to help reach optimal levels.

Interesting research regarding a microfluidic chip that facilitates multiplex analysis on a single biopsy tissue sample was recently published in a Microsystems & Nanoengineering article from José Luis García-Cordero’s group at Unidad Monterrey.  The technique allows highly irregular surfaces to be simply examined without further modification.

The PDMS chip, shown in upper image (©Nature – Microsystems & Nanoengineering), uses a ~3 x 7 mm chamber for the biopsy tissue sample, allows media to perfuse around the sample (orange fluid).  Once the valve is pneumatically actuated (7-35 kPa), the tissue is pressed against the lid to seal the microfluidic channels and allow the simultaneous introduction of discrete drug or tissue culture solutions through each of the eight channels.  Characterisation studies shown in the lower image (©Nature – Microsystems & Nanoengineering) demonstrate good permanent sealing and no cross-talk between channels.  Also, the continuous periperfusion of media around the sample increased tissue cell viability over the first several hours in the microfluidic device vs. in a petri dish.

The authors discuss the possibility of using biopsy samples in devices to perform drug efficacy, safety and toxicology studies before clinical trials, as well as facilitating research in physiology, metabolism and tissue regeneration.