On-chip acidification rate measurements from single cardiac cells confined in sub-nanoliter volumes

Abstract

The metabolic activity of cells can be monitored by measuring the pH in the extracellular environment. Microfabrication and microfluidic technologies allow the sensor size and the extracellular volumes to be comparable to single cells. A glass substrate with thin film pH sensitive IrO x electrodes was sealed to a replica-molded polydimethylsiloxane (PDMS) microfluidic network with integrated valves. The device, termed NanoPhysiometer, allows the trapping of single cardiac myocytes and the measurement of the pH in a detection volume of 0.36 nL. For wild-type (WT) single cardiac myocytes an acidification rate of 6.45 ± 0.38 mpH/min was measured in comparison to 19.5 ± 0.38 mpH/min for very long chain Acyl-CoA dehydrogenase (VLCAD) deficient mice in 0.8 mM of Ca 2+ . VLCAD deficiency is a fatty acid oxidation disease leading to cardiomyopathy and arrhythmias. The acidification rate increased to 11.96 ± 1.33 mpH/min for WT and to 32.0 ± 4.64 mpH/min for VLCAD −/− in 1.8 mM of Ca 2+ . The NanoPhysiometer concept can be extended to study ischemia/reperfusion injury or disorders of other biological systems to identify strategies for treatment and possible pharmacological targets.

DNA-barcode directed capture and electrochemical metabolic analysis of single mammalian cells on a microelectrode array

“…In earlier work, Castellarnau et al used dielectrophoresis to localize high concentration suspensions of bacteria near an ISFET pH sensor and measured the acidification of the cells in the presence of glucose.29 While this technique was well suited to measurement of the bulk response, it lacks the ability to resolve the unique activity of single cells. The single cardiac cell pH system of Ges et al 15 provides the ability to monitor large adherent cells, but the volume displacement caused by sealing the channel makes it difficult to direct the cell attachment. DNA-barcode capture provides the advantage of directed capture of both adherent and naturally non-adherent cells, such as T and B cells.…”

“…Ges et al recently demonstrated a device for on-chip measurement of acidification rates from single cardiac myocytes using physical confinement. 15 In their system, single myocytes were isolated in the sensing volume by physically pinching closed the ends of a PDMS channel. While this system represents an important step in single cell monitoring, the cell isolation technique does not allow for controlled capture on the sensor electrodes, which would be necessary for spatially resolved multi-analyte monitoring from single cells.…”

Heart-on-a-chip based on stem cell biology

“…Heart models in microsystems presented in literature allow research on the single cells level (Kaneko et al, 2007, Ges et al, 2008, Murthy et al, 2006, Cheng et al, 2006, multicellular cultures (Nguyen et al, 2009, Nguyen et al, 2015, Pong et al, 2011, Annabi et al, 2013, Ren et al, 2013, Grosberg et al, 2011 and 3D tissue tests (Huang et al, 2007b, Horiguchi et al, 2009, Cheah et al, 2010, Chen et al, 2013 (Table 1). These microsystems are more representative experimental models of the in vivo environment, than conventional culture dishes.…”

Towards monitoring real-time cellular response using an integrated microfluidics-matrix assisted laser desorption ionisation/nanoelectrospray ionisation-ion mobility-mass spectrometry platform

“…Extending our existing μL-scale electrochemical technologies [15,16,17,18], we are now combining at the nL scale optical and electrochemical technologies to record dynamic changes in acidification rate, oxygen and glucose consumption, lactate release, intracellular calcium concentrations, transmembrane potentials, and NADH concentrations. Sensitivities approaching that required to monitor single cells have been achieved in preliminary work [19,20,21,22,23,24]. Together these technologies are ideal for studying paracrine and autocrine signaling dynamics.…”