3. Multi-parametric monitoring of 3D cultures
Impedance changes are a compilation of multiple processes occurring in
cell culture systems. In the case of 2D cultures, significant impedance
changes have been successfully correlated with cell proliferation and
death. However, in 3D cultures, cellular processes are more complex,
thus impedance changes may be influenced by different factors such as
matrix degradation, cell spreading, migration or proliferation. This
raises the need for a secondary or multi‑parametric approach to further
understand, monitor and validate 3D culture systems. In‑line monitoring
systems coupled with impedance can lead to better real-time and
multi‑parametric biological understanding of complex 3D systems. Zhang
et al. developed a multisensor-integrated organ-on-chip platform that
contains two organ modules, one physical/chemical sensing module and one
bioelectrochemical sensing module, all connected via a perfused
system.(Zhang et al., 2017) The physical/chemical sensing system
consisted of optical pH and oxygen sensors. Additionally, gold
electrodes functionalized with desired antibodies for specific detection
of molecules in the media could be incorporated into the
bioelectrochemical sensing module. The EIS changed upon binding of the
target molecule and detection of different concentrations of the
analyte/biomarker was possible. They found that the amount of biomarker
that could be captured by the functionalized electrode surface was
proportional to the concentration in the solution, with detection levels
of 0.1 to 100 ng/ml, similar to many commercial ELISA systems. Thus,
biologically relevant concentrations were detected. Even though this is
a complex system, it contains fundamental parameters for real-time
monitoring of organoids. However, this system is highly modular and
specific to the microfluidics system they utilize. This makes it harder
to standardize and integrate these systems with current tissue culture
consumables and microscopes. Nevertheless, to date, this is the most
complete multi-parametric monitoring system for organoids.
Others have also used a multi-parametric monitoring approach coupled
with electrical measurements to monitor 3D cell cultures. Misun and
colleagues correlated amperometric (current) changes to glucose and
lactate concentrations in spheroids, giving real-time information on the
metabolic state of these microtissues. For the detection of glucose and
lactose in these spheroids, a microfluidic polydimethylsiloxane (PDMS)
chip with an enzyme-based hydrogel coated onto sensing electrodes was
proposed.(Misun, Hierlemann, & Frey, 2018) However, the sensor chip and
well was limited in its use as it is specifically designed for spheroid
growth via the hanging-drop culture technique. The sensor also depends
on the effective release of glucose from the spheroid to the media.
Another approach to monitor glucose concentrations and cell
proliferation was shown by Curto et al. They developed an in vitro cell
monitoring platform, which monitored the impedance changes of tissue by
means of an organic electrochemical transistor (OECT) at the bottom of
the well.(Vincenzo F. Curto et al., 2017) Glucose was also monitored
with an OECT-based glucose biosensor, but the glucose biosensor was not
in situ. Media was collected at the outlet of the perfused system and
later measured with the sensor. This multi parameter system allowed the
monitoring of kidney cell metabolic state via the glucose sensor as well
as changes in resistance and capacitance in real-time while the kidney
cell epithelium recovered from an electrical wound.
All of the above systems allow for real-time monitoring of the metabolic
activity of cells through measuring glucose, pH and oxygen levels in the
media. Even though they are sensitive, they depend on the efficient
release of the molecule of interest into the media, which can be limited
in a 3D cell culture system. The lack of spatial resolution displayed by
most of the monitoring techniques within these devices remains a
drawback. Correlating metabolite levels in media with a technique that
gives information about spatial resolution would allow for more
specialized biological information to be gained from the monitoring
system.