1 research outputs found
Open-Channel Microfluidic Membrane Device for Long-Term FT-IR Spectromicroscopy of Live Adherent Cells
Spatially resolved infrared spectroscopy
is a label-free and nondestructive
analytical technique that can provide spatiotemporal information on
functional groups in biomolecules of a sample by their characteristic
vibrational modes. One difficulty in performing long-term FT-IR measurements
on live cells is the competition between the strong IR absorption
from water and the need to supply nutrients and remove waste. In this
proof of principle study, we developed an open-channel membrane device
that allows long-term continuous IR measurement of live, adherent
mammalian cells. Composed of a gold-coated porous membrane between
a feeding channel and a viewing chamber, it allows cells to be maintained
on the upper membrane surface in a thin layer of fluid while media
is replenished from the feeding channel below. Using this device,
we monitored the spatiotemporal chemical changes in living colonies
of PC12 cells under nerve growth factor (NGF) stimulation for up to
7 days using both conventional globar and high-resolution synchrotron
radiation-based IR sources. We identified the primary chemical change
cells undergo is an increase in glycogen that may be associated with
secretion of glycoprotein to protect the cells from evaporative stress
at the air–liquid interface. Analyzing the spectral maps with
multivariate methods of hierarchical cluster analysis (HCA) and principal
component analysis (PCA), we found that the cells at the boundary
of the colony and in a localized region in the center of the colony
tend to produce more glycogen and glycoprotein than cells located
elsewhere in the colony and that the degree of spatial heterogeneity
decreases with time. This method provides a promising approach for
long-term live-cell spectromicroscopy on mammalian cell systems