ECM558 QCM™ Tumor Cell Transendothelial Migration Assay (Colorimetric, 24 Assays)

ECM558
1 kit  24 Assays
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      Overview

      Replacement Information

      Key Specifications Table

      Detection Methods
      Colorimetric
      Description
      Catalogue NumberECM558
      Trade Name
      • QCM
      • Chemicon
      DescriptionQCM™ Tumor Cell Transendothelial Migration Assay (Colorimetric, 24 Assays)
      OverviewAlso available: Cell Comb™ Scratch Assay! Get biochemical data from a scratch assay! Click Here

      Introduction
      A quantitative assay for tumor transendothelial migration has been described using a modified Boyden chamber system, (Okada, 1994; Li, 1999; Laferriere, 2001). The Boyden Chamber system is a two-chamber system with a porous membrane providing an interface between these two chambers. Endothelial cells are cultured on top of the porous membrane that is coated with an extracelluar matrix (ECM) protein. A tumor cell suspension is added above the endothelial monolayer. The invasion of tumor cells across the endothelium is determined by measuring the number of cells that migrate to the lower chamber.

      Millipore's QCM Tumor Cell Transendothelial Cell Migration Assay - Colorimetric provides an efficient model to analyze the ability of tumor cells to invade the endothelium. The assay is designed with an 8 µm pore size cell culture insert, appropriate for most cancer cell lines. The upper side of the cell culture insert is coated with fibronectin to support the optimal attachment and growth of endothelial cells. The assay allows investigators to compare the invasiveness of a variety of tumor cell lines, and to evaluate the effects of various factors influencing the process.

      Precoated cell culture inserts are provided in the Millipore QCM Tumor Cell Transendothelial Cell Migration Assay to significantly reduce assay time. Additionally, the assay allows quantitative analysis of tumor cell migration. Following incubation of tumor cells with the endothelial cell layer, invasive tumor cells are stained and quantified. In a departure from traditional Boyden methodology, stain is eluted with extraction buffer, transferred to a microplate, and measured spectrophotometrically. (Prior to elution, the investigator has the option of counting cells individually, if desired.) Spectrophotometric absorbance correlates with cell migration.
      Background InformationTumor metastasis is a multistep cascade. In order for tumor cells to migrate from a primary tumor mass to distant locations, they must invade through the basal membrane and into blood vessels (intravasation), circulate in the blood stream, survive during transport, then migrate out of a blood vessel (extravasation) to establish micrometastases. The penetration of circulating tumor cells into the endothelium is a crucial step for tumor metastasis.
      Materials Required but Not Delivered1. Endothelial cells (for example: HUVECs), and cell culture medium (for example: EGM-2)
      2. Invasive tumor cell lines and cell culture medium
      3. Harvesting buffer: Millipore's cell detachment solution, Accutase™ (Cat. No. SCR005), EDTA or trypsin-based cell detachment buffer, or other cell detachment formulations as optimized by individual investigators can also be used
      4. Serum-free medium, such as DMEM, MEM, etc. containing 0.5% BSA
      5. Sterile PBS or HBSS to wash cells
      6. Distilled water
      7. (Optional) Chemoattractant or pharmacological agent for addition to culture medium
      8. Low speed centrifuge and tubes for cell harvesting
      9. CO2 incubator appropriate for subject cells
      10. Hemocytometer or other means of counting cells
      11. Trypan blue or equivalent viability stain
      12. Microplate reader (540-570 nm detection) or spectrophotometer
      13. (Optional) Graduated ocular (calibrated), or automated method for counting stained cells on a membrane

      References
      Product Information
      Components
      • 1. Migration Test Plate: (Part No. CS202627) Two 24-well culture plates, each containing 12 human fibronectin-coated cell culture inserts (8 μm pore size), sufficient for the evaluation of 24 test samples
      • 2. TNFα: (Part No. 2004167) One tube - 20 μL at 0.1 mg/mL.
      • 3. Cell Stain Solution*: (Part No. 20294) One vial - 10 mL
      • 4. Extraction Buffer: (Part No. 20295) One vial - 10 mL
      • 5. 24 well Stain Extraction Plate: (Part No. 2005871) One plate.
      • 6. 96 well Stain Quantitation Plate: (Part No. 2005870) One plate.
      • 7. Swabs: (Part No. 10202) 50 each.
      • 8. Forceps: (Part No. 10203) 1 pair.
      Detection methodColorimetric
      Applications
      ApplicationThis QCM Tumor Cell Transendothelial Cell Migration Assay -Colorimetric provides an efficient model to analyze the ability of tumor cells to invade the endothelium.
      Application NotesCalculation of Results

      Results of the assay may be illustrated graphically using a bar chart to display optical density (OD) values at ~570 nm.

      A typical tumor cell transendothelial migration experiment will compare with a negative control. Negative control chambers, containing HUVECs only, function to determine the level of HUVECs migrating through the membrane. Cell migration in these chambers is generally low as there is no ECM on the lower side of the membrane to support endothelial migration, and these chambers are typically used as "blanks" for interpretation of data. As such, migration in test wells can be described as the value of tumor transendothelial migration minus the amount of migration visualized in the HUVEC only control.

      Additional migration may also be induced or inhibited in test wells through the addition of cytokines or other pharmacological agents.
      The following figures demonstrate typical migration results. One should use the data at left for reference only. This data should not be used to interpret actual assay results.
      Biological Information
      Physicochemical Information
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Usage Statement
      • Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
      Storage and Shipping Information
      Storage ConditionsTNFα should be stored at -20°C. All other components should be store at 2° to 8°C. Please refer to kit label for expiration date.

      Packaging Information
      Material Size1 kit
      Material Package24 Assays
      Transport Information
      Supplemental Information
      Specifications

      Documentation

      QCM™ Tumor Cell Transendothelial Migration Assay (Colorimetric, 24 Assays) SDS

      Title

      Safety Data Sheet (SDS) 

      References

      Reference overviewPub Med ID
      Transendothelial migration of colon carcinoma cells requires expression of E-selectin by endothelial cells and activation of stress-activated protein kinase-2 (SAPK2/p38) in the tumor cells.
      Laferriere, J, et al.
      J. Biol. Chem., 276: 33762-72 (2001)  2001

      Show Abstract
      11448946 11448946
      A modified Boyden chamber assay for tumor cell transendothelial migration in vitro.
      Li, Y H and Zhu, C
      Clin. Exp. Metastasis, 17: 423-9 (1999)  1999

      10651309 10651309
      A novel in vitro assay system for transendothelial tumor cell invasion: significance of E-selectin and alpha 3 integrin in the transendothelial invasion by HT1080 fibrosarcoma cells.
      Okada, T, et al.
      Clin. Exp. Metastasis, 12: 305-14 (1994)  1994

      7518760 7518760

      User Guides

      Title
      QCM Tumor Cell Transendothelial Migration Assay - Colorimetric

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      Categories

      Life Science Research > Cell Analysis > Cell-based Assays > Cell Migration Assays