Measuring Cell-Edge Protrusion Dynamics during Spreading using Live-Cell Microscopy

Nikola Lukic; Trishna Saha; Stefanie Lapetina; Michal Gendler; Gilad Lehmann; Anthony J. Koleske; Hava Gil-Henn

doi:10.3791/63157

Measuring Cell-Edge Protrusion Dynamics during Spreading using Live-Cell Microscopy

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Biology

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JoVE Journal Biology

Measuring Cell-Edge Protrusion Dynamics during Spreading using Live-Cell Microscopy

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05:50 min

November 1, 2021

DOI: 10.3791/63157-v

Nikola Lukic*¹, Trishna Saha*¹, Stefanie Lapetina¹, Michal Gendler¹, Gilad Lehmann¹, Anthony J. Koleske^2,3, Hava Gil-Henn¹

¹The Azrieli Faculty of Medicine,Bar-Ilan University, ²Department of Molecular Biophysics and Biochemistry,Yale University, ³Department of Neuroscience,Yale University

Overview

This study presents a cell-edge protrusion assay to investigate the dynamic parameters of spreading cells, including protrusions, retractions, and ruffles. This method is particularly significant as it correlates with cell migration, aiding in the identification of critical proteins and signaling mechanisms involved in cell motility.

Key Study Components

Research Area

Cell migration
Cell motility
Cytoskeletal dynamics

Background

A correlation exists between cell-edge protrusions and cell migration.
The assay is straightforward, cost-effective, and does not necessitate fluorescent labeling.
Understanding protrusions can provide insights into cellular behavior and mechanisms driving motility.

Methods Used

Cell-edge protrusion assay
Adhered cells on a glass bottom dish
Kymography analysis for assessing protrusions

Main Results

Average frequency of protrusions was 5.1 per 10 minutes, with ruffles at 2.1.
Kymography analysis revealed a protrusion distance of approximately 4.8 micrometers.
Identified the importance of choosing appropriate cells in the spreading phase for accurate results.

Conclusions

The study introduces an accessible assay to assess cell motility dynamics.
This method serves as a preliminary assessment tool before more complex migration assays are undertaken.

Frequently Asked Questions

What is the main purpose of the cell-edge protrusion assay?

The assay helps measure the dynamic parameters of cell motility, like protrusions and retractions.

Is fluorescent labeling required for this method?

No, the method is designed to be simple and cost-effective without the need for fluorescent labeling.

What are the critical metrics analyzed in this study?

Protrusions, retractions, and ruffles are analyzed to understand cell dynamics.

How does the assay contribute to biology research?

It aids in identifying proteins and signaling mechanisms involved in cell migration and motility.

What temperature conditions are required for cell incubation?

Cells should be incubated at 37 degrees Celsius during the assay.

Are there specific conditions for choosing cells for imaging?

Yes, cells must be in their spreading phase and not in contact with other cells to avoid interference.

What is the significance of kymography in this research?

Kymography is used to visualize and quantify the dynamics of the protrusions over time.

This protocol aims to measure the dynamic parameters (protrusions, retractions, ruffles) of protrusions at the edge of spreading cells.

The cell-edge protrusion assay has been shown to directly correlate with cell migration. Therefore it can be used as a preliminary method for identifying critical proteins and signaling mechanisms involved in cell motility. The method is fast, simple, cost effective and does not require fluorescent labeling or using an expensive fluorescent microscope.

Demonstrating the procedure will be Michal Gendler, a student from my laboratory. Add two milliliters of one normal hydrochloric acid solution at the center of a glass bottom dish and incubate for 20 minutes at room temperature. Wash the dish three times with two milliliters of PBS.

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