Cultrex Laminin I Cell Invasion Assay, 96-well Summary
• Standardized assay for Laminin I cell invasion
• Available in 24 and 96 well formats
• Allows optimization for specific cell lines
Why Use Cultrex Laminin I Cell Invasion Assay, 96-well?
The Cultrex Laminin I Cell Invasion Assays provide systems where the researcher can determine the optimal Laminin I coating for their research. Since different cell lines and different treatments can result in a wide range of invasive potentials, the permissiveness of each cell line to invade through Laminin I may be optimized to fit each experiment by adjusting the coating concentration. The Cultrex Laminin I Cell Invasion Assays are provided as either a single 96 well plate providing capacity for large screening experiments or as 24 individual inserts providing flexibility for smaller studies.
Cultrex Mouse Laminin I is purified from Engelbreth-Holm-Swarm (EHS) tumor. Laminin I is a major component of the basement membrane which is a continuous sheet of specialized extracellular matrix that form an interface between endothelial, epithelial, muscle, or neuronal cells and their adjacent stroma and that play an essential role in tissue organization by influencing cell adhesion, migration, proliferation, and differentiation.
• 10X Coating Buffer
• 5X Laminin I Solution
• Calcein AM
• Cell Invasion Chamber
• 25X Cell Wash Buffer
• 10X Cell Dissociation Solution
Note: The components for this kit may require different storage/shipping temperatures and may arrive in separate packaging.
For research use only. Not for diagnositic use.
FBS Stimulates Migration of NIH-3T3 and HT1080 Cells. The NIH-3T3 mouse embryonic fibroblast cell line and the HT1080 human fibrosarcoma cell line were treated with 10% fetal bovine serum (FBS). The migration of untreated (yellow bars) and treated (green bars) NIH-3T3 and HT1080 cells against different extracellular matrix components, including Cultrex BME, Laminin I, Collagen I, Collagen IV, were quantified using the Cultrex Cell Invasion Assay Kits (Catalog # 3455-096-K, 3456-096-K, 3457-096-K, 3458-096-K, respectively). Data from four experiments was quantified for both non-invasive (NIH-3T3) and invasive (HT1080) cell types.
What is cell invasion?
Cell invasion is cell migration through a physiological barrier in response to a chemoattractant, and this recapitulates cell movement within a physiological environment which is composed of extracellular matrix proteins. Cultrex® Cell Invasion Assays evaluate cell invasion based on the cells ability to traverse membranes that are coated with a layer of extracellular matrix proteins. The cells must traverse this barrier through a combination of protein degradation and cellular locomotion.
What are the variables associated with cell invasion?
The major variables associated with cell invasion are cell type, cell density, composition of the physiological barrier, thickness of the physiological barrier, chemoattractant that is used, and time of culture.
Will the Cultrex® Cell Invasion Assay work for my cells?
The Cutltrex Cell Invasion Assay is currently configured for invasive adherent cell lines. If your cell line is adherent and there is evidence in the scientific literature that your cell line is invasive in a Boyden chamber, it should be compatible with our assay. If this is unknown, then the invasive potential will need to be determined empirically.
How should cells be cultured prior to setting up the Cultrex Cell Invasion Assay?
Cells need to be healthy and actively dividing in 2-D culture. Cells should be passaged two or three times after resuspension from cryopreservation, and they should never surpass 80% confluency during each passage. Cells should also be assessed for viability using trypan blue, and they should exhibit less than 5% staining.
How do the Cultrex Cell Invasion Assays compare to wound healing assays?
Wound healing assays, also known as scratch assays, monitor cell migration laterally on a tissue culture treated plate. This is accomplished by generating a void in a cell monolayer by either removing cells or treating the surface of the plate to prevent cell growth in a designated area. The assay measures the ability of the cell monolayer to fill this void, and it may be conducted in the presence of extracellular matrix proteins. Since this assay is conducted within one chamber, there is no chemotactic gradient, and without the membrane, the cells are no longer required to change shape and squeeze through the pores. Another potential problem is that this assay does not control for differences in cell proliferation. While wound healing assays may be valuable for supplementing the Boyden chamber assay, it is not a replacement.
Can the Calcein-labeled invasive cells be subcultured?
Calcein AM cytotoxicity should be determined empirically for each cell line or model. For best results, the cells should be removed from the cell dissociation solution and placed in fresh culture medium as soon as possible.
The kit comes with both black and clear microplates. Do I need to use both?
The entire assay may be conducted in either the clear or black assay plate (provided in3455-096-01), if desired. The black assay plate provides increased sensitivity and reduced background for plate readers that read from the top. The clear assay plate provides compatibility for plate readers that read from the bottom. Some researchers have used the clear assay plate for the invasion step and the black assay plate for detection (top read).
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