Functional Phase Imaging

Figure 1.  The combined optical setup for simultaneous quantitative phase imaging and fluorescence imaging. The fluorescence excitation, fluorescence emission and phase imaging wavelengths are all spectrally separated using dichroic mirrors.
Figure 2. Quantitative phase imaging (QPI) and fluorescence (or Förster) resonance energy transfer (FRET) imaging of caspase-3 mediated apoptosis. (A) QPI and (B) FRET images of HeLa cells undergoing apoptosis over time. Black-dotted square indicates QPI field of view (FOV) relative to the FRET FOV (C) Acceptor and donor average emission over time showing clear demonstration of diminished FRET signal in the cell indicated by the white arrow. (D) FRET ratio and optical volume (OV) over time for same cell in (C). A rapid OV loss was seen approximately 21 minutes after caspase-3 activation as indicated by the drop in FRET ratio. All scale bars are microns.

Chemical, mechanical, and electrical stimuli have all been shown to modulate the transmission state of active ion channels in cells. However, there is still a lack of understanding of how mechanical and electrical stimuli interact to produce changes in cell signaling pathways and sub-cellular structures. Figure 1 shows our custom-built microscopy system capable of performing simultaneous quantitative phase microscopy (QPM) and Förster resonance energy transfer (FRET) measurement to study the structural, mechanical, and electrical properties of cells at the nanoscale. An example of this capability is seen in Figure 2, where we are tracking the optical volume and FRET signal of cells undergoing apoptosis. FRET imaging can be exploited to assess local tension through nanoscale measurements of force transduction by specific molecules within cell structures. FRET sensors can also be utilized to detect both membrane potential and concentration of calcium ions to relate mechanical information to channel activation. Integration of QPM allows for visualization of cell structure with nanometer depth sensitivity and at millisecond time scales. Previously, we developed QPM methods to use this nanoscale information to distinguish cell phenotype based on mechanical properties, track stiffness changes during carcinogenesis, and estimate shear moduli from our QPM shear flow assay. We seek to further develop this QPM toolbox and use it in conjunction with FRET to study how external stimuli are transduced by cellular structures to produce changes in the internal electrical state via modulation of ion channel transmission. Fig 2. from Eldridge et. al. 2018.

Associated Lab Members


  • Silvia Ceballos
  • Will Eldridge
  • Michael Habib
  • Latifah Maasarani
  • Han Sang Park
  • Steven Parker
  • Albert Rancu