Label-free Live Cell Imaging & Analysis
The HoloMonitor® label-free live cell imaging system is based on the principle of quantitative phase imaging, enabling non-invasive visualization and live cell analysis of cultured cells without compromising cell integrity.
How Does It Work?
Just like water waves, light waves of a specific wavelength have two principal characteristics: amplitude and phase. Amplitude corresponds to light intensity and is the height of the wave, measured from crest to trough. Phase describes whether a wave is currently at its crest, in its trough, or somewhere in between.
For a living cell to be visible to the naked eye or in a light microscope, the light arriving from the cell must differ in intensity from the background, i.e. differ in amplitude. Unfortunately, living cells are as translucent as ice cubes in water. They neither absorb, emit nor scatter light to any significant extent and therefore only slightly change the amplitude of the illuminating light, if at all.
Phase-shifted beach waves.
The Live Cell Imaging Problem
To be visible in a light microscope, cells must be stained to absorb, emit or scatter light. Unstained living cells do, however, slow down and distort the light passing through them, just like beach waves are distorted by shallower water (above).
By using a phase contrast microscope these distortions or phase-shifts can be observed, making unstained cells clearly visible. However, conventional live cell imaging using phase contrast microscopy cannot quantify phase-shifts, only visualize them.
The Live Cell Analysis Solution
With computer technology, it is possible both to quantify and visualize phase-shifts. This technique is called quantitative phase imaging (QPI) or quantitative phase contrast microscopy to distinguish it from conventional phase contrast microscopy. QPI provides both quantitative and beautiful images of living cells, transforming phase microscopy and live cell imaging into a quantitative tool for detailed live cell analysis.
An example of a quantitative phase image of living cells in 3D, created by HoloMonitor. The height of the cell and its color tone correspond to the optical thickness of the cell.
Live Cell Imaging References
Evaluation of Holographic Imaging Cytometer HoloMonitor M4 Motility ApplicationsCytometry Part A (2018)Read more
The HoloMonitor software modules for cell tracking and wound healing analysis were evaluated and compared to the more conventional methods transwell migration and transwell invasion. Both HoloMonitor modules were found to be well-correlated with established standards, yielded reproducible results, and at the same time offered distinct advantages. The wound healing assay was the most tractable and automated method with good reproducibility, while the cell tracking module enabled identification of hypermobile subpopulations.
Label-free High Temporal Resolution Assessment of Cell Proliferation Using Digital Holographic MicroscopyCytometry Part A (2017)Read more
The authors have developed a robust and label-free kinetic cell proliferation assay with high temporal resolution for adherent cells using HoloMonitor M4. Only two image processing settings were adjusted between cell lines, making the assay practical, user friendly, and free of user bias. In the recorded time-lapse image sequences, individual cells were automatically identified to provide detailed growth curves and growth rate data of cell number, confluence, and average cell volume. The results demonstrate how these parameters facilitate a deeper understanding of cell processes than what is achievable with current single-parameter and end-point methods.
HoloMonitor M4: holographic imaging cytometer for real-time kinetic label-free live-cell analysis of adherent cellsProceedings, Quantitative Phase Imaging II (2016)Read more
Live-cell imaging enables studying dynamic cellular processes that cannot be visualized in fixed-cell assays. An increasing number of scientists in academia and the pharmaceutical industry are choosing live-cell analysis over or in addition to traditional fixed-cell assays. We have developed a time-lapse label-free imaging cytometer HoloMonitor M4. HoloMonitor M4 assists researchers to overcome inherent disadvantages of fluorescent analysis, specifically effects of chemical labels or genetic modifications which can alter cellular behavior. Additionally, label-free analysis is simple and eliminates the costs associated with staining procedures.