Reproducibility of scratch assays is affected by the initial degree of confluence: Experiments, modelling and model selection

Wang, Jin; Shah, Esha; Penington, Catherine J.; McCue, Scott W.; Chopin, Lisa K.; Simpson, Matthew J.

doi:10.1016/j.jtbi.2015.10.040

Cited by 124 publications

(424 citation statements)

References 34 publications

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“…The differences how cells proliferate in the scratch assay and the proliferation assay is surprising because we are making observations well away from the location of the scratch. This finding that we have two phases of proliferation in scratch assays is significant because many mathematical studies implicitly assume that cells in scratch assays proliferate logistically for the entire duration of the experiment [14,49,54,76,77,98,100,118]. However, our finding is that cells located far away from the scratch proliferate very differently to cells in the proliferation assay.…”

Section: Introductionmentioning

confidence: 59%

“…To provide a more thorough investigation of the suitability of various mathematical models, we calibrate mathematical models to a suite of experimental data where the initial density of cells is intentionally varied [49]. To achieve this, our experimental procedure involves placing a different number of cells into each well of the tissue culture plate.…”

Section: Initial Cell Densitymentioning

confidence: 99%

“…In addition, the results from our previous study, focusing on scratch assays, suggest that when calibrating solutions of a logistic-type reaction-diffusion equation to experimental data with varying initial cell density, there appears to be no unique value of λ for which the logistic growth equation matches the entire data set for all initial cell densities [49]. One way of interpreting this result is that the cells in the scratch assay do not proliferate logistically.…”

Section: Introductionmentioning

confidence: 97%

“…In the applied mathematics literature, scratch assays have been modelled using continuum reaction-diffusion equations [14,49,54,76,77,98,100,102,103,106]. In these models, cell migration is represented by a diffusion term, and carrying-capacity limited proliferation is represented by a logistic source term.…”

Section: Introductionmentioning

confidence: 99%

“…It is interesting to note that a logistic growth term is often used when modelling scratch assays or proliferation assays [14,49,54,76,77,98,100,118], yet the suitability of this choice is rarely, if ever, tested using experimental data. In fact, several studies argue that the logistic growth equation does not always match experimental data [66,97,124,130].…”

Section: Introductionmentioning

confidence: 99%

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Investigating the Reproducibility of In Vitro Cell Biology Assays Using Mathematical Models

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Section: Introductionmentioning

confidence: 59%

Section: Initial Cell Densitymentioning

confidence: 99%

Section: Introductionmentioning

confidence: 97%

Section: Introductionmentioning

confidence: 99%

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confidence: 99%

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Investigating the Reproducibility of In Vitro Cell Biology Assays Using Mathematical Models

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Label‐Free Cell Migration Assay Using Magnetic Exclusion

Gupta

Dabaghi

et al. 2021

Adv Materials Technologies

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biophysical cues. [1,2] Signaling molecules regulate cell migration under normal physiological conditions by controlling cell adhesion and cytoskeleton organization. [1,3,4] Conversely, pathological conditions such as metastatic cancer, vascular disease, and chronic inflammation are associated with unregulated cell migration, contributing to disease progression. [4,5] The wound healing assay is a commonly used in vitro method to study cell migration, cell interactions, and the underlying mechanisms. [6][7][8][9][10] The conventional "scratch" assay creates a cellfree area or scratch in a cell monolayer using a pipette tip or a pin tool. [10,11] Cells from intact surrounding regions migrate and repopulate the cell-free region over time. The migration rate is measured by observing the decrease in the cellfree area at regular intervals. While the scratch assay is simple and inexpensive, it requires 24-48 h to form a cell monolayer. [11] Moreover, by physically scratching the cell layer with a pipette tip or microneedle, cell-free regions of inconsistent shapes and sizes are created. [10,12,13] This process also damages the underlying plastic surface or extracellular matrix (ECM) substrate, which can significantly affect cell migration. [10,12,13] The electric cell-substrate impedance sensing (ECIS) technique, on the other hand, produces consistent cell-free regions without mechanically disrupting the cell layer. [10,12] In ECIS, a confluent cell layer is formed over a small gold film electrode placed at the bottom of a tissue culture well. Electric pulses are applied to induce cell death at the electrode surface, which creates a well-defined region devoid of cells. [14] However, dead cell fragments can remain on top of the electrode and interfere with cell migration. [12,15] Furthermore, the electric pulses can potentially injure the cells surrounding the electrode, thereby affecting their migration. [10] In contrast to the scratch assay and ECIS, the physical exclusion technique creates cell-free areas of reproducible sizes and shapes with minimal cellular damage. [12,16] This method employs physical barriers, such as plastic inserts or silicone stoppers, to prevent cells from settling in a predefined area. However, it requires 12-20 h to form confluent cellular regions around the barrier. [16,17] Commercial inserts are also expensive and unsuitable for multiple uses. [16] Magnetic levitation is another in vitro technique used to form cell clusters in a migration assay. [18] This method is based on the cellular uptake of biopolymer-encapsulated magnetic In vitro wound healing assays are widely used to investigate cell migration during various physiologic and pathologic processes. However, traditional scratch-based assays produce cell-free areas that are not reproducible, whereas the alternate insert-based exclusion method is expensive and timeconsuming. Here, a rapid, label-free, insert-free magnetic exclusion technique, where magnetic fields are used to create cell-free areas is described. Suspensions of diamagnet...

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Non-local physics informed neural networks for forward and inverse problems containing non-local operators

Singh,

Mehra,

Pulch

2024

Neural Comput & Applic

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Reproducibility of scratch assays is affected by the initial degree of confluence: Experiments, modelling and model selection

Cited by 124 publications

References 34 publications

Investigating the Reproducibility of In Vitro Cell Biology Assays Using Mathematical Models

Investigating the Reproducibility of In Vitro Cell Biology Assays Using Mathematical Models

Label‐Free Cell Migration Assay Using Magnetic Exclusion

Non-local physics informed neural networks for forward and inverse problems containing non-local operators

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