In real-time, the HoloMonitor® Cell Proliferation Imaging Assay automatically provides growth curves of adherent cells by cell count and confluence, inside your incubator.
Cell proliferation assays are widely applied in biomedical sciences, especially for potency and cytotoxicity testing of drugs and chemicals. The assays are commonly used to understand how drug dose levels affect cells over time, typically by creating cell growth curves to determine the inhibitory concentration at half maximum effect (IC50).
To assess cell growth at various treatment concentrations, the number of cells in multiple samples with increasing treatment concentrations are traditionally counted each day over several days, using time-consuming manual cell counting. Automated cell counters have made cell counting less labor-intensive, but the cells still need to be destructively prepared and transferred to the cell counter.
To make cell proliferation assessment more convenient, rapid chemometric assays have been developed, such as MTT, XTT, AlamarBlue and Neutral Red. These indirect assays measure metabolic activity or other cellular properties that are assumed to correspond to the number of cells.
By repeatedly counting the number of cells in a cell culture, automated image‐based methods provide direct and hence more relevant assessment of cell proliferation than the widely used chemometric methods.
However, to be a viable option, time-lapse‐based adherent cell counting must be label‐free and fully automated — not relying on manual or user-aided cell identification and segmentation. Furthermore, to be compatible with established laboratory workflow, it is vital that any novel cell proliferation assay can be robustly performed using standard cell culture microplates.
Janicke et al., Label‐free high temporal resolution assessment of cell proliferation using digital holographic microscopy, Cytometry Part A (2017)
For various microplate formats, the HoloMonitor Cell Proliferation Assay conveniently records non-invasive quantitative time-lapse images of multiple cell cultures inside your incubator. While the time-lapse image sequences are automatically recorded, the assay presents real-time growth curves by cell count and cell confluence at user-selected intervals.
When using the HoloMonitor Cell Proliferation Assay, cell count and confluence growth curves are updated and displayed in real-time throughout the experiment.
As cell proliferation assessment using HoloMonitor is entirely non-invasive and does not require assay-specific sample preparation, the recorded time-lapse image sequence can be reused to determine other cellular characteristics associated with potency and cytotoxicity. Using, for example, the HoloMonitor Cell Morphology Assay, Cell Motility Assay and the HoloMonitor Dose Response Assay, cell morphology, motility and dose response can, in addition, be quantified without requiring additional experiments, saving precious cells and lab hours.
The utility of this method is that it is quick, quantitative, and qualitative, which is essential for future cell-based research.
Digital Holographic Cytometry: Macrophage Uptake of Nanoprobes, Imaging & Microscopy (2019)
The HoloMonitor® Live Cell Assays
(screenshot of the HoloMonitor App Suite software)
After the experiment, cell proliferation data is easily exported to Excel.
HoloMonitor video of proliferating cells, showing frequent cell divisions.
Various nanoprobes are designed to have drug-delivery or detection properties. However, as a foreign bodies in human blood circulation, nanoprobes can potentially induce inflamatory response and active macrophages. In this study, L. Sternbæk et al. have employed HoloMonitor M4 to study the effects of nanoprobes (molecularly imprinted polymers targeted to sialic acid) on macrophage cell line, RAW 264.7 by evaluating cell morphology (cell area, volume, and thickness) and cell count. Authors have shown that neither the HoloMonitor M4 nor nanoprobes affected the physiological functions of macrophages.
Review describing and comparing various assays used to study biocompatibility and cytotoxicity of nanomaterials. Holographic phase imaging is pointed out as an excellent tool for cell morphometric characterisation and cell migration studies, and the authors conclude that the interest in the use of DH microscopy in research is constantly increasing.
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.
In this PHI webinar, our application specialist Lisa Bodily introduces the exciting field of label-free live cell imaging. We discuss live cell imaging techniques and highlight reasons to work label-free. You will learn how digital holographic microscopy visualizes your cells and get to know our HoloMonitor system. Also, our label-free applications for cell biology research are presented and some examples highlighted.
View the webinar on demand here