Evaluation of the Cell Invasion and Migration Process: A Comparison of the Video Microscope-based Scratch Wound Assay and the Boyden Chamber Assay
Jean‐Baptiste GuySophie EspenelAlexis VallardPriscillia Battiston-MontagneAnne‐Sophie WoznyDominique ArdailGersende AlphonseC. RancouleClaire Rodriguez‐LafrasseNicolas Magne
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The invasion and migration abilities of tumor cells are main contributors to cancer progression and recurrence. Many studies have explored the migration and invasion abilities to understand how cancer cells disseminate, with the aim of developing new treatment strategies. Analysis of the cellular and molecular basis of these abilities has led to the characterization of cell mobility and the physicochemical properties of the cytoskeleton and cellular microenvironment. For many years, the Boyden chamber assay and the scratch wound assay have been the standard techniques to study cell invasion and migration. However, these two techniques have limitations. The Boyden chamber assay is difficult and time consuming, and the scratch wound assay has low reproducibility. Development of modern technologies, especially in microscopy, has increased the reproducibility of the scratch wound assay. Using powerful analysis systems, an "in-incubator" video microscope can be used to provide automatic and real-time analysis of cell migration and invasion. The aim of this paper is to report and compare the two assays used to study cell invasion and migration: the Boyden chamber assay and an optimized in vitro video microscope-based scratch wound assay.Keywords:
Gentamicin protection assay
Scratch
Video microscopy
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The invasion and migration abilities of tumor cells are main contributors to cancer progression and recurrence. Many studies have explored the migration and invasion abilities to understand how cancer cells disseminate, with the aim of developing new treatment strategies. Analysis of the cellular and molecular basis of these abilities has led to the characterization of cell mobility and the physicochemical properties of the cytoskeleton and cellular microenvironment. For many years, the Boyden chamber assay and the scratch wound assay have been the standard techniques to study cell invasion and migration. However, these two techniques have limitations. The Boyden chamber assay is difficult and time consuming, and the scratch wound assay has low reproducibility. Development of modern technologies, especially in microscopy, has increased the reproducibility of the scratch wound assay. Using powerful analysis systems, an "in-incubator" video microscope can be used to provide automatic and real-time analysis of cell migration and invasion. The aim of this paper is to report and compare the two assays used to study cell invasion and migration: the Boyden chamber assay and an optimized in vitro video microscope-based scratch wound assay.
Gentamicin protection assay
Scratch
Video microscopy
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The invasion and migration abilities of tumor cells are main contributors to cancer progression and recurrence. Many studies have explored the migration and invasion abilities to understand how cancer cells disseminate, with the aim of developing new treatment strategies. Analysis of the cellular and molecular basis of these abilities has led to the characterization of cell mobility and the physicochemical properties of the cytoskeleton and cellular microenvironment. For many years, the Boyden chamber assay and the scratch wound assay have been the standard techniques to study cell invasion and migration. However, these two techniques have limitations. The Boyden chamber assay is difficult and time consuming, and the scratch wound assay has low reproducibility. Development of modern technologies, especially in microscopy, has increased the reproducibility of the scratch wound assay. Using powerful analysis systems, an "in-incubator" video microscope can be used to provide automatic and real-time analysis of cell migration and invasion. The aim of this paper is to report and compare the two assays used to study cell invasion and migration: the Boyden chamber assay and an optimized in vitro video microscope-based scratch wound assay.
Scratch
Video microscopy
Gentamicin protection assay
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Several common wound healing models have been used to evaluate wound healing agents and formulations, namely: conditioned media (CM), transwell co-cultures (TWCC) and co-cultures (CC) in a monolayer. However, no study has been conducted to compare the relevance of these models in the keratinocytes and fibroblasts interaction physiologically. Therefore, this study aimed to compare these models based on cell migration and proliferation, and matrix metalloproteinase (MMP) expression.Cell migration was analysed by scratch assay and MMP-7, while cell proliferation was analysed by (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) tetrazolium reduction assay.Increased cell migration was observed in CM and TWCC models, while varied results were obtained in CC. Cell migration was increased due to upregulation of MMP-7 in CM and TWCC models, while it was downregulated in CC, which might have hindered migration of both cells in monolayers.CM and TWCC are more suitable than CC for wound healing research and for evaluating wound healing agents or formulations, as they can better simulate the layered tissue constructs and paracrine interactions in the physiological environment.
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The wound-healing assay is simple, inexpensive, and one of the earliest developed methods to study directional cell migration in vitro. This method mimics cell migration during wound healing in vivo. The basic steps involve creating a "wound" in a cell monolayer, capturing the images at the beginning and at regular intervals during cell migration to close the wound, and comparing the images to quantify the migration rate of the cells. It is particularly suitable for studies on the effects of cell-matrix and cell-cell interactions on cell migration. A variation of this method that tracks the migration of individual cells in the leading edge of the wound is also described in this chapter.
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Wound healing is a complex event involving both cellular and molecular activities. Fibroblasts play an important role and are keys to wound healing through cell proliferation and migration. Honey has anti-microbial, anti-oxidant, anti-inflammatory properties, which are used for various benefits such as wound healing. This study aims to explore the effect of honey on the viability and migration ability of fibroblast cells. The method used is the viability test using the MTT Assay calculated by the formula for the percentage of cell viability. Migration test using In Vitro Wound Scratch Assay. The results of the migration test images were analyzed using ImageJ. Giving honey doses of 0.5% and 0.1% increased cell viability and migration after 24 hours of intervention. Decreased cell viability after 48 hours of treatment, but there was a difference in the meaning of honey 1%, 0.5%, and 0.1% compared to control. Honey doses of 1%, 0.5%, and 0.1% increased fibroblast cell migration compared to control. The lowest honey increases the viability and migration of fibroblasts so that the possibility of wound healing.
Keywords: Honey; Fibroblast Migration; Wound Scratch Assay; Wound Healing
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Cell migration is a dynamic process, which is important for embryonic development, tissue repair, immune system function, and tumor invasion 1, 2. During directional migration, cells move rapidly in response to an extracellular chemotactic signal, or in response to intrinsic cues 3 provided by the basic motility machinery. Random migration occurs when a cell possesses low intrinsic directionality, allowing the cells to explore their local environment. Cell migration is a complex process, in the initial response cell undergoes polarization and extends protrusions in the direction of migration 2. Traditional methods to measure migration such as the Boyden chamber migration assay is an easy method to measure chemotaxis in vitro, which allows measuring migration as an end point result. However, this approach neither allows measurement of individual migration parameters, nor does it allow to visualization of morphological changes that cell undergoes during migration. Here, we present a method that allows us to monitor migrating cells in real time using video - time lapse microscopy. Since cell migration and invasion are hallmarks of cancer, this method will be applicable in studying cancer cell migration and invasion in vitro. Random migration of platelets has been considered as one of the parameters of platelet function 4, hence this method could also be helpful in studying platelet functions. This assay has the advantage of being rapid, reliable, reproducible, and does not require optimization of cell numbers. In order to maintain physiologically suitable conditions for cells, the microscope is equipped with CO2 supply and temperature thermostat. Cell movement is monitored by taking pictures using a camera fitted to the microscope at regular intervals. Cell migration can be calculated by measuring average speed and average displacement, which is calculated by Slidebook software.
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Wound healing is a dynamic and complex process of replacing missing or dead cell structures and tissue layers. The aim of this research is to discover biocompatible materials and drugs that can promote cell migration in the wound area and thus enhance desirable wound healing effects. In this paper, we report that PDMS nanogratings could accelerate the migration of epithelial cells along the grating axis, and the addition of Imatinib could further increase the epithelial cell wound healing speed to 1.6 times the speed of control cells. We also demonstrate that this migration is mediated by lamellipodia protrusion, and is Rac1-GTPase activity dependent. Lastly, we discuss the potential application and prospect of different nanostructured biomaterials for wound healing studies.
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Cell migration is a dynamic process, which is important for embryonic development, tissue repair, immune system function, and tumor invasion 1, 2. During directional migration, cells move rapidly in response to an extracellular chemotactic signal, or in response to intrinsic cues 3 provided by the basic motility machinery. Random migration occurs when a cell possesses low intrinsic directionality, allowing the cells to explore their local environment. Cell migration is a complex process, in the initial response cell undergoes polarization and extends protrusions in the direction of migration 2. Traditional methods to measure migration such as the Boyden chamber migration assay is an easy method to measure chemotaxis in vitro, which allows measuring migration as an end point result. However, this approach neither allows measurement of individual migration parameters, nor does it allow to visualization of morphological changes that cell undergoes during migration. Here, we present a method that allows us to monitor migrating cells in real time using video - time lapse microscopy. Since cell migration and invasion are hallmarks of cancer, this method will be applicable in studying cancer cell migration and invasion in vitro. Random migration of platelets has been considered as one of the parameters of platelet function 4, hence this method could also be helpful in studying platelet functions. This assay has the advantage of being rapid, reliable, reproducible, and does not require optimization of cell numbers. In order to maintain physiologically suitable conditions for cells, the microscope is equipped with CO2 supply and temperature thermostat. Cell movement is monitored by taking pictures using a camera fitted to the microscope at regular intervals. Cell migration can be calculated by measuring average speed and average displacement, which is calculated by Slidebook software.
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