Chemotaxis and migration tool 2

Manufactured by Ibidi

Sourced in Germany, United States

The Chemotaxis and Migration Tool 2.0 is a lab equipment designed for the study of cell migration and chemotaxis. It provides a controlled and standardized environment for researchers to observe and analyze the movement of cells in response to chemical gradients or other stimuli.

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Lab products found in correlation

Axio observer z1, by Zeiss (3 mentions) Zen 2, by Zeiss (2 mentions) Axiocam 503 mono camera, by Zeiss (2 mentions) Ec plan neofluar 10 0.30 ph1 air objective, by Zeiss (2 mentions) Culture insert, by Ibidi (2 mentions) Six well plate, by Corning (2 mentions) Laminin, by R&D Systems (1 mentions) Transwell chambers 3 μm pore, by Corning (1 mentions) Eclipse ni microscope, by Nikon (1 mentions) Glass bottom petri dishes, by MatTek (1 mentions) Two well culture insert, by Ibidi (1 mentions) Axio observer z1 inverted, by Zeiss (1 mentions) Prism 7, by GraphPad (1 mentions) Tcs sp8 microscope, by Leica (1 mentions) Biostation im q, by Nikon (1 mentions) Nis elements advanced research, by Nikon (1 mentions) Biostation ct, by Nikon (1 mentions) Imagexpress pico system, by Molecular Devices (1 mentions) Fty720, by MedChemExpress (1 mentions) Vivaview incubator fluorescence microscope, by Olympus (1 mentions)

20 protocols using chemotaxis and migration tool 2

Evaluating Chemotaxis and Cell Motility

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Cell motility parameters of maximum Euclidean and Accumulated distances and COM vector resolution in the Y-axis direction for each cell tracked over 48 hrs, were resolved via sequential use of Nd-to-Image6d (National Institutes of Health, Bethesda, USA), Manual Tracking (Fabrice Cordelières, Institut Curie, Orsay, France), and Chemotaxis and Migration Tool 2.0 (ibidi, Munich, Germany) plug-ins, all running on an ImageJ platform as previously described97 (link). COM is a strong parameter for evaluating chemotaxis, and measures the spatial average displacement of all cell endpoints with positive or negative coordinates, depending on the direction of movement of a single cell or a population of cells. COM displacement in the Y-axis further specifies directionality of cell movement towards the SRR. Cell tracking data selected for analysis included only video recordings of RPC and PPC movement between 18 and 42 hrs to account for complete 24 hr periods of sustained steady-state gradients of SDF-1α within the microchannel. A two-sample Student’s t-test (t) assuming unequal variances was performed to assess significant difference in mean motility parameters between control and test groups.

Unachukwu U.J., Warren A., Li Z., Mishra S., Zhou J., Sauane M., Lim H., Vazquez M, & Redenti S. (2016). Predicted molecular signaling guiding photoreceptor cell migration following transplantation into damaged retina. Scientific Reports, 6, 22392.

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Directed Migration of Cells in 2D Assay

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For the 2D assay, cell culture inserts and the Chemotaxis and Migration Tool 2.0 (Ibidi, Madison, WI) were used. Inserts were coated with 20 μg/ml laminin (R&D Systems). PKH26-red-fluorescent-ECs in EGM media were plated on one side of the insert, and GFP-CSCs in complete NBM on the other side. Subsequently, the media was replaced with complete NBM, the insert removed, and live-imaging of cells into the gap initiated using a 10X objective lens and a motorized-stage (Leica-DMI6000-ImageEM/Orca-R2-ImageProPlus). Manual Tracking (Image J) was used to trace EC paths into the 500 μm gap [60 (link)]. ECs trajectories were projected onto the XY plane and the average velocity calculated for individual and total trajectories [60 (link)].
For analysis of CSCs/ECs migrating into the wound gap, an automated algorithm was generated by ImageIQ (Cleveland, OH) designed to batch process and analyze time-lapse, multi-channel stacks within ImagePro-Plus-7.0.
For the Transwell^® assay, cells were seeded on the upper filter surface (Transwell chambers, 3-μm pore, Corning), allowed to migrate, cells on the upper surface removed, and cells on the lower filter surface washed, fixed [11 (link)], photographed and counted in 5-10 fields with a 20X objective (Leica-DFC425C-QImaging-Q15729-QCapturePro).

Burgett M.E., Lathia J.D., Roth P., Nowacki A.S., Galileo D.S., Pugacheva E., Huang P., Vasanji A., Li M., Byzova T., Mikkelsen T., Bao S., Rich J.N., Weller M, & Gladson C.L. (2016). Direct contact with perivascular tumor cells enhances integrin αvβ3 signaling and migration of endothelial cells. Oncotarget, 7(28), 43852-43867.

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Quantifying Directional Cell Migration

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Wilcoxon, Mann-Whitney U test, Friedmann and Kurskal-Wallis-Test combined with Dunn’s post hoc test were used for statistical analysis as indicated. Levels of significance were labeled as follows: *p < 0.1, **p < 0.01, ***p < 0.001, ****p < 0.0001. Directionality was assessed by dividing the euclidian distance (shortest distance between initial and final position in a straight line) and the accumulated distance of the total cell migration path. Analyses were performed and graphs were designed using GraphPad Prism (GraphPad Software, Inc., San Diego, California) and BioRender (Toronto, Canada). Polar Plots were created using the Chemotaxis and Migration Tool 2.0 (IBIDI, Martinsried, Germany) and Fiji.

Preuß S.L., Oehrl S., Zhang H., Döbel T., Engel U., Young J.L., Spatz J.P, & Schäkel K. (2023). Immune complex-induced haptokinesis in human non-classical monocytes. Frontiers in Immunology, 14, 1078241.

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Quantifying Cell Migration in Collagen Gels

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The cell migration in time-lapse images was manually tracked using NIH ImageJ software (National Institutes of Health, Bethesda, MD, USA). The cell tracking data were further quantified using the Chemotaxis and Migration Tool 2.0 (ibidi USA, Fitchburg, WI, USA). Cell migration velocity and distance were analyzed in collagen gels with different concentrations. For cell migration under EFs, the method was previously reported [26 (link)]. The cell migration direction was expressed as the cosine of the angle. The directedness of each experimental group was calculated from the equation

(\sum i c o s θ) / n

, where

n

is the cell number of all replicates. The cell migration velocity was calculated from the accumulated cell migration distance during the recording time.

Yao L., Tran K, & Nguyen D. (2022). Collagen Matrices Mediate Glioma Cell Migration Induced by an Electrical Signal. Gels, 8(9), 545.

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Visualizing Tracheal Development and bnl Expression

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Live and fixed cell imaging were performed using either a spinning disc confocal microscope (Andor and Perkin Elmer) or a scanning confocal microscope (Leica SP5X). For time lapse imaging of the developing trachea and bnl source, dechorionated embryos were imaged through glass bottom petri dishes (MatTek), over 60 μm z-stacks, every 10 minutes for a duration of ∼5-6 hours in an Andor spinning disc microscope. Cells in embryo were tracked using the Tracking plugin in Fiji. Subsequent analyses of migration of the dorsal- and lateral- bnl-expressing clusters were carried out with Chemotaxis and Migration Tool 2.0 from ibidi. In situ hybridization images were acquired with an upright Nikon Eclipse Ni microscope and a color CCD camera. The images were analyzed with iQ3 and Fiji software.

Du L., Zhou A., Patel A., Rao M., Anderson K, & Roy S. (2017). Generation of a targeted expression system for branchless and characterization of novel cellular expression patterns of the gene in Drosophila. Developmental biology, 427(1), 35-48.

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Wound Healing and Cell Migration Assay

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Wound healing and open-field migration assays were performed using a two-well culture insert (Ibidi). pHCAECs were resuspended in 70 μl of complete vascular growth medium and plated at a density of 1,100 cells per well into each reservoir of a culture insert (Ibidi) that adhered to the bottom of a six-well plate (Corning) and grown in standard cell culture conditions. After confluence is reached, the insert was removed using forceps, and the plate was rinsed with DPBS to remove cell debris. Migrating cells were cultured in complete vascular growth medium supplemented or not with 100 ng/ml rh TNFα. Time-lapse images were acquired, at 10-min intervals over 24 h, in standard cell culture conditions on an Axio Observer Z1 inverted motorized microscope (Zeiss) equipped with an incubation system for live-cell imaging (PeCon), using an EC Plan-Neofluar × 10/0.30 Ph1 air objective, Axiocam 503 mono camera, and ZEN 2 software (all from Zeiss). Phase contrast images were recorded at a location of the migration gap and sites of open-field migration. Cell movement was analyzed using the Manual Tracking plugin for ImageJ (NIH). To quantify the dynamics of cell migration, such as velocity or Euclidean distance and accumulated distance, the migration trajectories were then assessed with Chemotaxis and Migration Tool 2.0 (Ibidi).

Gagat M., Zielińska W., Mikołajczyk K., Zabrzyński J., Krajewski A., Klimaszewska-Wiśniewska A., Grzanka D, & Grzanka A. (2021). CRISPR-Based Activation of Endogenous Expression of TPM1 Inhibits Inflammatory Response of Primary Human Coronary Artery Endothelial and Smooth Muscle Cells Induced by Recombinant Human Tumor Necrosis Factor α. Frontiers in Cell and Developmental Biology, 9, 668032.

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Confluent Cell Migration Assay

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Confluent cell migration assay was performed using six-well plates (Corning). After confluence is reached, the cells were cultured in complete vascular growth medium supplemented or not with 100 ng/ml rh TNFα. Time-lapse images were acquired, at 10-min intervals over 36 h in standard cell culture conditions on an Axio Observer Z1 inverted motorized microscope (Zeiss) equipped with an incubation system for live-cell imaging (PeCon), using an EC Plan-Neofluar × 10/0.30 Ph1 air objective, Axiocam 503 mono camera, and ZEN 2 software (all from Zeiss). Phase contrast images were recorded at a location of the migration gap and sites of open-field migration. Cell movement was analyzed using the Manual Tracking plugin for ImageJ (NIH). To quantify the dynamics of cell migration, such as velocity or Euclidean distance and accumulated distance, the migration trajectories were then assessed with Chemotaxis and Migration Tool 2.0 (Ibidi).

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Cell Migration Statistical Analysis

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The significant differences between two groups were calculated using an unpaired t-test or one-way ANOVA and Kruskal–Wallis test with Dunn’s correction for multiple comparisons. Relations between cell migration parameters were assessed using Pearson’s correlation coefficient analysis. Statistical analyses were performed by using Prism 7 software (GraphPad). The Rayleigh test was used to determine cell movement homogeneity using Chemotaxis and Migration Tool 2.0 (Ibidi). A p-value < 0.05 was considered to be statistically significant and labeled on figures as ^∗ or $, p < 0.05; ^∗∗ or

$ $

, p < 0.01; ^∗∗∗ or

$ $ $

, p < 0.001; ^**** or

$ $ $ $

, p < 0.0001; and NS, non-significant. Results were expressed as means ± SD.

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Scratch Wound Assay for Cell Migration

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Confluent monolayers of cells were transfected with the siRNA or plasmid of the indicated gene for 24 h, and a scratch wound model then was created. To inhibit cell proliferation, the serum in the medium changed from 10 to 2%. Cells were then placed on the stage of a Leica TCS SP8 microscope (Wetzlar, Germany) in a humidified atmosphere (at 37°C, 5% CO₂) and observed with a 10× lens. Images were acquired every 40 min and analyzed using ImageJ software (National Institutes of Health). Trajectory analysis was performed using the Chemotaxis and Migration Tool 2.0 (Ibidi, GmbH, München, Germany).

Qu H., Miao T., Wang Y., Tan L., Huang B., Zhang L., Liu X., Long M., Zhang R., Liao X., Gong X., Wang J., Xiong X., Liu J., Li X., Yu J., Yang G., Zhu Z., Zheng H, & Zheng Y. (2021). Dedicator of Cytokinesis 5 Regulates Keratinocyte Function and Promotes Diabetic Wound Healing. Diabetes, 70(5), 1170-1184.

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Tracking Cell Motility under Graphene Exposure

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The freshly harvested control and graphene-treated A549 cells were seeded into the 35 mm dish with a polymer coverslip bottom. The seeding density was 1 × 105 cells per dish in 2 mL of the DMEM culture medium supplemented with 10% fetal bovine serum. The cells were allowed to recover overnight, and the dish was then placed into the Biostation IM-Q (Nikon Instruments, Inc., Melville, NY, USA). The time-lapse video of the cells was recorded, with cells captured every 15 min for a total of 24 h. Cell motility was evaluated using the MTrackJ manual tracking plugin of the FIJI software [22 (link)]. A total of 100 control or graphene-treated cells were tracked. The mean velocity and the mean accumulated distance were calculated using the Chemotaxis and Migration Tool plugin for the FIJI Software (Chemotaxis and Migration Tool 2.0, Ibidi, Gräfelfing, Germany, free download from http://www.ibidi.de/applications/ap_chemo.html accessed on 12 July 2020).

Šestáková B., Schröterová L., Bezrouk A., Čížková D., Elkalaf M., Havelek R., Rudolf E, & Králová V. (2022). The Effect of Chronic Exposure of Graphene Nanoplates on the Viability and Motility of A549 Cells. Nanomaterials, 12(12), 2074.

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