Dataset Information

Significantly improved precision of cell migration analysis in time-lapse video microscopy through use of a fully automated tracking system.

ABSTRACT:

Background

Cell motility is a critical parameter in many physiological as well as pathophysiological processes. In time-lapse video microscopy, manual cell tracking remains the most common method of analyzing migratory behavior of cell populations. In addition to being labor-intensive, this method is susceptible to user-dependent errors regarding the selection of "representative" subsets of cells and manual determination of precise cell positions.

Results

We have quantitatively analyzed these error sources, demonstrating that manual cell tracking of pancreatic cancer cells lead to mis-calculation of migration rates of up to 410%. In order to provide for objective measurements of cell migration rates, we have employed multi-target tracking technologies commonly used in radar applications to develop fully automated cell identification and tracking system suitable for high throughput screening of video sequences of unstained living cells.

Conclusion

We demonstrate that our automatic multi target tracking system identifies cell objects, follows individual cells and computes migration rates with high precision, clearly outperforming manual procedures.

SUBMITTER: Huth J

PROVIDER: S-EPMC2858025 | biostudies-literature | 2010 Apr

REPOSITORIES: biostudies-literature

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Publications

Significantly improved precision of cell migration analysis in time-lapse video microscopy through use of a fully automated tracking system.

Huth Johannes J Buchholz Malte M Kraus Johann M JM Schmucker Martin M von Wichert Götz G Krndija Denis D Seufferlein Thomas T Gress Thomas M TM Kestler Hans A HA

BMC cell biology 20100408

<h4>Background</h4>Cell motility is a critical parameter in many physiological as well as pathophysiological processes. In time-lapse video microscopy, manual cell tracking remains the most common method of analyzing migratory behavior of cell populations. In addition to being labor-intensive, this method is susceptible to user-dependent errors regarding the selection of "representative" subsets of cells and manual determination of precise cell positions.<h4>Results</h4>We have quantitatively an ...[more]

PMID: 20377897

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Project description:Time-lapse imaging of cell colonies in microfluidic chambers provides time series of bioimages, i.e., biomovies. They show the behavior of cells over time under controlled conditions. One of the main remaining bottlenecks in this area of research is the analysis of experimental data and the extraction of cell growth characteristics, such as lineage information. The extraction of the cell line by human observers is time-consuming and error-prone. Previously proposed methods often fail because of their reliance on the accurate detection of a single cell, which is not possible for high density, high diversity of cell shapes and numbers, and high-resolution images with high noise. Our task is to characterize subpopulations in biomovies. In order to shift the analysis of the data from individual cell level to cellular groups with similar fluorescence or even subpopulations, we propose to represent the cells by two new abstractions: the particle and the patch. We use a three-step framework: preprocessing, particle tracking, and construction of the patch lineage. First, preprocessing improves the signal-to-noise ratio and spatially aligns the biomovie frames. Second, cell sampling is performed by assuming particles, which represent a part of a cell, cell or group of contiguous cells in space. Particle analysis includes the following: particle tracking, trajectory linking, filtering, and color information, respectively. Particle tracking consists of following the spatiotemporal position of a particle and gives rise to coherent particle trajectories over time. Typical tracking problems may occur (e.g., appearance or disappearance of cells, spurious artifacts). They are effectively processed using trajectory linking and filtering. Third, the construction of the patch lineage consists in joining particle trajectories that share common attributes (i.e., proximity and fluorescence intensity) and feature common ancestry. This step is based on patch finding, patching trajectory propagation, patch splitting, and patch merging. The main idea is to group together the trajectories of particles in order to gain spatial coherence. The final result of CYCASP is the complete graph of the patch lineage. Finally, the graph encodes the temporal and spatial coherence of the development of cellular colonies. We present results showing a computation time of less than 5?min for biomovies and simulated films. The method, presented here, allowed for the separation of colonies into subpopulations and allowed us to interpret the growth of colonies in a timely manner.

Project description:Quantifying and understanding movement is critical for a wide range of questions in basic and applied ecology. Movement ecology is also fostered by technological advances that allow automated tracking for a wide range of animal species. However, for aquatic macroinvertebrates, such detailed methods do not yet exist. We developed a video tracking method for two different species of benthic macroinvertebrates, the crawling isopod Asellus aquaticus and the swimming fresh water amphipod Gammarus pulex. We tested the effects of different light sources and marking techniques on their movement behavior to establish the possibilities and limitations of the experimental protocol and to ensure that the basic handling of test specimens would not bias conclusions drawn from movement path analyses. To demonstrate the versatility of our method, we studied the influence of varying population densities on different movement parameters related to resting behavior, directionality, and step lengths. We found that our method allows studying species with different modes of dispersal and under different conditions. For example, we found that gammarids spend more time moving at higher population densities, while asellids rest more under similar conditions. At the same time, in response to higher densities, gammarids mostly decreased average step lengths, whereas asellids did not. Gammarids, however, were also more sensitive to general handling and marking than asellids. Our protocol for marking and video tracking can be easily adopted for other species of aquatic macroinvertebrates or testing conditions, for example, presence or absence of food sources, shelter, or predator cues. Nevertheless, limitations with regard to the marking protocol, material, and a species' physical build need to be considered and tested before a wider application, particularly for swimming species. Data obtained with this approach can deepen the understanding of population dynamics on larger spatial scales and of the effects of different management strategies on a species' dispersal potential.

Dataset Information

Significantly improved precision of cell migration analysis in time-lapse video microscopy through use of a fully automated tracking system.

Background

Results

Conclusion

Publications

Significantly improved precision of cell migration analysis in time-lapse video microscopy through use of a fully automated tracking system.

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