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Label-Free Tracking of Adherent Cells

The HoloMonitor® Adherent Cell Tracking facilitates label-free live cell time-lapse imaging and characterization of hetero­geneous cell behavior on a single-cell level.

Tracking Heterogeneous Cell Behavior

Mammalian cells can be highly dynamic in both morphology and behavior. Characterizing and tracking heterogeneous cell behavior over time on a single-cell level is critically important when studying rare events, such as the acquisition of therapeutic resistance, or transition events, such as differentiation.

The Limitations of Labeled Tracking

Quantitative live cell imaging using high content analysis allows for kinetic evaluation of adherent cells. However, fluorescent labels are often required for accurate cell state classification. The disruptive and cytotoxic effects commonly associated with fluorescent dyes can limit the length of time single cells are tracked unperturbed. Additionally, fluorescent labels must be applied a priori to classify cell states of interest, despite observations that a single gene expression is often insufficient to predict behavior or cell state.

Individual Cell Behavior Matters

Furthermore, it is increasingly acknowledged that the outcome of cancer therapy is determined by the response of individual cells on a molecular level. Live cell time-lapse imaging and single-cell tracking is a powerful tool to understand the behavior of individual cells and to gain insight into how cell-to-cell variability translates to the overall drug response of an entire cell population.

Label-Free Cell Tracking

HoloMonitor facilitates this growing demand for label-free kinetic classification of subpopulations within heterogeneous unperturbed cell cultures.

The possibility to track adherent cells is the most advanced and powerful feature of HoloMonitor time-lapse cytometer. It allows cell movement and morphology of adherent cells to be monitored and analyzed over extended periods of time, without requiring labels nor genetic modifications. Motion and kinetic morphology data are provided individually for each cell and as mean values for all cells included in the analysis.

label-free adherent cell tracking

The HoloMonitor® Live Cell Assays
(screenshot of the HoloMonitor App Suite software)

Our [single-cell tracking] data clearly show that there are sub­populations of rapidly dividing cells, hiding in population-based data such as the population doubling time. Thus, by only drawing conclusions based on population data, important biological processes on individual cell levels may be neglected.

Kamlund et al.

Single-Cell Motility vs. Migration

Cell movement is categorized both by non-directional cell motility and by directional cell migration. Cell motility is the random cell movement, occurring in varying magnitude in almost every cell culture. Cell migration is the non-random movement caused by a cell attractant or repellent.

label-free adherent single-cell tracking, cell motility

A spatial plot created by the HoloMonitor Adherent Cell Tracking software, showing that one cell stands out among the other cells by having a clear directional movement.

Video showing L929 cells, first untreated and thereafter treated with colcemid. Notice how the chemotherapy drug dramatically reduces cell motility.

Kinetic Cell Morphology

In addition to cell motion, HoloMonitor quantifies a wide range of morphological properties of tracked cells, for example, cell volume, area, thickness, irregularity and roughness. Each of these properties may be plotted as time series to observe and quantify how they vary over time and over cell divisions.

label-free single-cell tracking, cell volume

In the above time-series, the cell volume of a single cell is tracked during its cell division, showing a steep increase in volume before division followed by an immediate halving of the cell volume when the cell has divided.

Excel Export

If preferred, all live cell tracking data can easily be exported to Excel for further analysis and graphical presentation.

Cell Tracking for Cell Morphology and Movement Analysis

The HoloMonitor® Adherent Cell Tracking offers robust, non-invasive kinetic quantification of cell morphology and movement.

  • Automated acquisition of multiple time-lapse image sequences in multiple wells.
  • Robust image segmentation and tracking of cells of interest.
  • Time-series plots of a range of morphological properties, revealing changes in individual cell morphology.
  • Single-cell motility and cell migration measurements.
  • Directness, i.e. the ratio of migration and motility, indicating how direct a cell or a sub­population moves.
  • The non-invasive nature of the method allows subsequent cellular staining for further analysis.

Key Cell Tracking Publications

  • Moving into a New Dimension: Tracking Migrating Cells with Digital Holographic Cytometry in 3D
    A. Görloff Wingren
    Cytometry Part A (2018)

    Commentary article discussing the fundamental role of cell movement studies in cancer research. Advantages of the HoloMonitor Cell Tracking and Wound Healing Applications over the transwell migration and invasion assays are highlighted, and includes the possibility to use the cells in experiments for other purposes after completing the imaging. In addition to single cell tracking, the HoloMonitor technology also benefits from the fact that morphology analysis can be performed of each cell. The author concludes that this indeed opens up for almost unlimited possibilities to perform cell morphology analysis using this methodology, since each image is very rich in cellular information.

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  • Influence of salinomycin treatment on division and movement of individual cancer cells cultured in normoxia or hypoxia evaluated with time-lapse digital holographic microscopy
    S. Kamlund, D. Strand, B. Janicke, K. Alm, S Oredsson
    Cell Cycle (2017)

    Digital holographic microscopy was used for longitudinal cell tracking of individual cells. Various parameters were investigated and the results suggest that EMT (epithelial to mesenchymal transision)  and MET (mesenchymal to epithelial transission) can be investigated using this method.

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  • High accuracy label-free classification of single-cell kinetic states from holographic cytometry of human melanoma cells
    M. Hejna et al.
    Scientific Reports (2017)

    The authors used singel-cell tracking and machine learning to develop a robust method for label-free classification of adherent cells.

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