Upper transwell chamber

Manufactured by Corning

Sourced in United States, Mexico, United Kingdom

The Upper Transwell Chamber is a component of a cell culture system used for studying cell migration and barrier function. It is a physical barrier that separates two compartments, allowing the exchange of soluble factors while restricting direct cell-cell contact.

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Lab products found in correlation

Matrigel, by BD (51 mentions) Matrigel, by Corning (25 mentions) Crystal violet, by Merck Group (13 mentions) Crystal violet, by Beyotime (11 mentions) Upper transwell chamber, by BD (8 mentions) Inverted microscope, by Olympus (7 mentions) Am6000 microscope, by Leica (7 mentions) Crystal violet, by Solarbio (6 mentions) Fetal bovine serum (fbs), by Thermo Fisher Scientific (5 mentions) Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (4 mentions) Inverted microscope, by Zeiss (4 mentions) Paraformaldehyde (pfa), by Beyotime (4 mentions) Paraformaldehyde (pfa), by Solarbio (3 mentions) Evos m7000, by Thermo Fisher Scientific (3 mentions) Matrigel, by Merck Group (3 mentions) Dulbecco modified eagle medium (dmem), by Corning (2 mentions) Type 1 collagen, by BD (2 mentions) Matrigel 1 15, by BD (2 mentions) Lower transwell chamber, by Corning (2 mentions) Optical microscope, by Olympus (2 mentions)

Close spelling variants of this product

24-well transwell upper chambers Gelatin-coated upper 8-µm pore transwell chambers Matrigel‐coated Transwell upper chambers Matrigel-coated upper chambers of Transwell inserts Non-coated upper chamber of transwell plates Upper chamber of transwell Upper transwell chamber insert Transwell upper chamber inserts Upper chamber Transwell chambers

223 protocols using upper transwell chamber

Matrigel-based Invasion Assay for Cell Migration

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The Matrigel-based invasion assay was performed as previously described [33 (link)]. Briefly, 2×10⁴ cells were plated in the upper Transwell chamber (Corning, NY, USA) in DMEM containing 1% FBS. The chamber basement membranewas coated with 60 μg Matrigel (Matrigel, Becton-DickinsonBiosciences, NJ). Then, the chamber was inserted into 24-well plates filled with 600 μL of DMEM medium containing 10% FBS. After incubation at 37°C for 24 h, the cells on the upper portion of the chamber membranes were removed, and the lower surfaces of the chamber membranes were fixed with 5% glutaraldehyde for 5 min and stained with 5% Giemsa. The number of invading cells from four randomly selected fields was counted under a microscope. For rescue assays, the A549 and H322 cells with stable expression of KLF17 protein were again transfected with pReceiver-M15-uPA containing uPA cDNA or a control vector; For effects of Src and p38/MAPK inhibition on invasion assay, the upper Transwell chamber (Corning, NY, USA) in DMEM containing 1% FBS and different concentration of SB 203580 (p38/MAPK inhibitor) or HY-13805 (PP2, Src inhibitor), and then the invasiveness of the cells was analyzed by Transwell assays in the same manner.

Cai X.D., Che L., Lin J.X., Huang S., Li J., Liu X.Y., Pan X.F., Wang Q.Q., Chen L., Lin M.J., Huang Z.H., Ma H.M., Wu Y., Liu S.M, & Zhou Y.B. (2017). Krüppel-like factor 17 inhibits urokinase plasminogen activator gene expression to suppress cell invasion through the Src/p38/MAPK signaling pathway in human lung adenocarcinoma. Oncotarget, 8(24), 38743-38754.

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Evaluating Cell Proliferation and Migration

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The cell proliferation capacity was evaluated using colony formation assay and cell counting kit 8 (CCK-8). Here, 2000 pretreated cells were seeded in each well of the 96-well plate for the CCK-8 assay. The absorbance value at 450 nm was determined after the addition of 10 μL CCK-8 reagent (MCE, NJ, USA) in each well for 1 h at 37 °C. As for the colony formation assay, 500 pretreated KIRC cells were seeded in each well of the six-well plate for 2 weeks. Relative colony rates were calculated after staining with 0.2% crystal violet and fixing in 4% paraformaldehyde. The cell migration and invasion abilities were measured using the transwell assay. Then, 200 μL serum-free DMEM medium with 2 × 104 pretreated cells were injected into the upper transwell chamber (Corning, New York, NY, USA) for migration assay. Similarly, cells were seeded in the upper transwell chamber precoated with Matrigel (Corning, New York, NY, USA) for invasion assay. As an attractant, 700 μL DMEM medium (10% FBS) was injected into the lower chamber. Migration or invasion assays were also subsequently treated with 4% paraformaldehyde and 0.2% crystal violet after 48 h.

Wu Y., Xu Z., Chen X., Fu G., Tian J., Shi Y., Sun J, & Jin B. (2023). Bioinformatics Prediction and Experimental Verification Identify CAB39L as a Diagnostic and Prognostic Biomarker of Kidney Renal Clear Cell Carcinoma. Medicina, 59(4), 716.

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Transwell Assay for Cell Migration and Invasion

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Transwell assay was applied to measure cell migration and invasion ability of NSCLC cells. For migration assay, a total of 4 × 10⁴ cells/well in medium supplemented with 5% FBS were placed in the upper transwell chamber (Corning, Cambridge, MA), medium containing 20% FBS was added to the lower chambers and cultured at 37°C, 5% CO₂ for 48 h. For invasion assay, cells were added to the upper transwell chambers, which were pre-coated with Matrigel. Other operations were the same as above. Then cells were fixed with 4% formaldehyde and stained with 0.1% Crystal Violet. The number of cells was counted under a microscope at 200× magnification.

Zhang W, & Feng G. (2021). C1QTNF6 regulates cell proliferation and apoptosis of NSCLC in vitro and in vivo. Bioscience Reports, 41(1), BSR20201541.

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Cell Migration and Invasion Assays

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For migration assay, 1 × 10⁵ 5637 cells or 4 × 10⁴ T24 cells were plated in the upper transwell chamber (Corning Ltd., USA) with 200‐μl serum‐free 1640 medium and 600 μl of 1640 medium containing 10% FBS in the lower chambers to induce cell migration. After incubated for 24 h at 37°C, cells were fixed with 4% paraformaldehyde and stained with .1% crystal violet. The migrated cell numbers were counted using phase contrast microscopy and statistically analysed.
For invasion assay, Matrigel was thawed and liquefied on ice and diluted in cold serum‐free 1640 medium to a final concentration of 200 μg/ml. Next, 100 μl of the diluted Matrigel was carefully added to the transwell insert and solidified in a 37°C incubator for 2 h to form a thin gel layer. Then, 1 × 10⁵ 5637 cells or 4 × 10⁴ T24 cells were plated in the upper transwell chamber (Corning Ltd., USA) with a 200‐μl‐serum‐free 1640 medium and 600 μl of 1640 medium containing 10% FBS in the lower chambers to induce cell migration. After incubated for 24 h at 37°C, cells were fixed with 4% paraformaldehyde and stained with .1% crystal violet. The migrated cell numbers were counted using phase contrast microscopy and statistically analysed.

Xiao Y., Ju L., Qian K., Jin W., Wang G., Zhao Y., Jiang W., Liu N., Wu K., Peng M., Cao R., Li S., Shi H., Gong Y., Zheng H., Liu T., Luo Y., Ma H., Chang L., Li G., Cao X., Tian Y., Xu Z., Yang Z., Shan L., Guo Z., Yao D., Zhou X., Chen X., Guo Z., Liu D., Xu S., Ji C., Yu F., Hong X., Luo J., Cao H., Zhang Y, & Wang X. (2022). Non‐invasive diagnosis and surveillance of bladder cancer with driver and passenger DNA methylation in a prospective cohort study. Clinical and Translational Medicine, 12(8), e1008.

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Cell Migratory and Invasive Potential

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Transwell migration and matrigel invasion assays were carried out utilizing Transwell chambers as indicated by the producer's guidelines. A matrigel grid was covered in the transwell membrane and utilized for the cell invasion test. In short, 1×10 5 cells/well in DMEM (100L,0.6%FBS) were put in the upper Transwell chamber (Corning Incorporated, NY, USA) that had been pre-coated with matrigel (Growth factor reduced, BD Biosciences, MD, USA). DMEM was loaded into the base chambers (20 % FBS). In the upper Transwell chamber, 1×10 5 cells/well in DMEM (100L, 0.5 % FBS) were placed for the migration test (Corning Incorporated, NY, USA).The base chambers were loaded up with DMEM (20% FBS). After 24 h, the cells were xed and stained in both the Transwell migration and matrigel invasion examine. Haphazardly chose elds were checked under a reversed magnifying lens (200 × magni cation, Carl ZEISS, Jena, German) and a normal worth was utilized as the quantity of attacked cells.

Dong L., Zheng J., Bai Z., Lu Y., Song W., Tang Y., & Linlin W. (2021). Upregulation of microRNA-483-5p Advances Non-small Cell Lung Cancer (NSCLC) Progression via Stifling HIPK2 Expression.

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Scratch and Transwell Migration Assays

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For the scratch migration assay, inducible U2OS cells were plated onto a 6-well plate and cultured to near (> 90%) confluence. Cells were serum starved for 24 h in the presence or absence of 2 μg/mL tetracycline, and the monolayer was scratched with a sterile 10 μL-pipette tip. Then serum-free media containing 10 ng/mL of TGF-β1 was added for 24 h. Phase-contrast images were acquired at 0 and 24 h after the gaps were created. The cells migrated into the gaps were counted from three different gap regions. For the transwell migration assay, transfected U2OS cells were resuspended in serum-free medium, and the cell suspension (4 × 10⁴ cells) was added to the upper transwell chamber (pore size of 8 μM; Costar; Corning, Corning, NY, USA). Media containing 0.1% serum and TGF-β1 was added to the bottom wells of the chambers. Cells were incubated for 18 h at 37°C, fixed with 4% paraformaldehyde, and permeabilized by 100% methanol. Cells were then stained with 0.5% crystal violet dissolved in 20% methanol at room temperature for 15 min. Cells that had not migrated after 18 h were removed from the upper face of the filters using cotton swabs. Migrated cells were counted under a light microscope. Images of three different fields were taken for each membrane.

Min K.W., Liggett J., Silva G., Wu W., Wang R., Shen R.F., Eling T, & Baek S. (2015). NAG-1/GDF15 Accumulates in the nucleus and modulates transcriptional regulation of the Smad pathway. Oncogene, 35(3), 377-388.

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Migration Assay for Cell Invasion

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For migration assay, 2.5 × 10⁵ cells suspended in 200 μl serum-free medium were added into the upper transwell chamber (8 mm, Corning Costar, USA), and 700 μl medium supplemented with 20% serum was placed in the lower chamber. Each transwell chamber contains a 6.5 mm diameter membrane with 8.0 μm pore size. After incubation, cells on the upper surface of membrane were removed gently with a cotton swab. Cells invading to the lower surface were fixed with methanol before staining with 0.1% crystal violet for 20 min at room temperature. The stained cells were counted in 5 randomly selected fields under a light microscope. Each clone was plated in triplicate in each experiment.

Zhang Y., Chen S., Zhu J., Guo S., Yue T., Xu H., Hu J., Huang Z., Chen Z., Wang P, & Liu Y. (2022). Overexpression of CBS/H2S inhibits proliferation and metastasis of colon cancer cells through downregulation of CD44. Cancer Cell International, 22, 85.

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Cell Migration and Invasion Assay

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For migration assay, 2 × 10⁴ cells suspended in 200 μl serum-free medium were added into the upper transwell chamber (8 mm, Corning Costar, USA), and 500 μl medium supplemented with 10% serum was placed in the bottom chamber. After 24 h incubation, cells on the upper surface were removed with a cotton swab. Cells invading to the lower surface of membrane were fixed with methanol before staining with 0.4% crystal violet for 30 min. The stained cells were counted in 5 randomly selected fields under an inverted microscope. The invasion assay was performed as above except that cells were plated on the chamber precoated with Matrigel (BD Bioscience, USA).

Wu H., Jing X., Cheng X., He Y., Hu L., Wu H., Ye F, & Zhao R. (2016). Asporin enhances colorectal cancer metastasis through activating the EGFR/Src/cortactin signaling pathway. Oncotarget, 7(45), 73402-73413.

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SIX3 and TRIM27 Regulation of Cell Migration

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Exponentially growing cells were seeded in 6-well plates with 300,000 cells per well and cultured overnight. A549 cells were subsequently infected with pLVX-Puro-SIX3 or blank vector, and H1975 cells were transfected with siSIX3 or siNC in the presence or absence of XAV939 (10 μM) or dimethyl sulfoxide (DMSO), otherwise H1975 cells were infected with pLVX-Puro-SIX3, pLVX-Puro-TRIM27, or blank vector in the presence or absence of XAV939 (10 μM) or DMSO, following culture overnight. Cells were then seeded in serum-free growth medium and incubated for one day. The upper transwell chamber (Costar, Cambridge, MA, USA) precoated with or without Matrigel (Becton Dickinson, Oxford, UK) was filled with cells resuspended in 300 μL RMPI-1640 containing 10% FBS with a density of 60,000 cells per well. An aliquot (700 μL) of RMPI-1640 containing 10% FBS was added to the lower chamber. After 24-h incubation, cells in the lower chamber were fixed with 4% paraformaldehyde and stained with crystal violet for 30 min. Non-migrated cells were removed with a cotton swab. Cell number was determined using a XDS-500D inverted microscope (Shanghai Caikon Optical Instrument Co., Ltd., Shanghai, China).

Liu S., Tian Y., Zheng Y., Cheng Y., Zhang D., Jiang J, & Li S. (2020). TRIM27 acts as an oncogene and regulates cell proliferation and metastasis in non-small cell lung cancer through SIX3-β-catenin signaling. Aging (Albany NY), 12(24), 25564-25580.

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Transwell Assay for VSMC Migration

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For the transwell assay, after the transfection of miR‐130a mimics, siRNA‐ATG2B or pCDNA3.1(+)‐ATG2B, VSMCs were resuspended in serum‐free DMEM (5 × 10⁵ cells/mL), and 200 µL of the solution was added to the upper transwell chamber (Costar, NY). The lower chamber was filled with DMEM (Gibco) supplemented with 10% FBS (Gibco). After 24 hours of incubation, the cells that migrated to the lower surface of the chamber membrane were fixed with paraformaldehyde (4%) and subsequently stained with 0.1% crystal violet in 20% methanol, whereas the cells that remained on the upper surface of the membrane were removed with a cotton swab. After 3 washes with PBS, the migrated VSMCs were counted under an inverted microscope (Leica DMI4000B).

Zheng L., Wang Z., Li Z., Wang M., Wang W, & Chang G. (2021). MicroRNA‐130a inhibits proliferation of vascular smooth muscle cells by suppressing autophagy via ATG2B. Journal of Cellular and Molecular Medicine, 25(8), 3829-3839.

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