Key Specifications Table
|Description||QCM™ Leukocyte Transendothelial Migration Assay (Colorimetric, 24 Assays)|
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Millipore’s Colorimetric QCM™ Leukocyte Transendothelial Cell Migration Assay provides an efficient model to analyze the ability of leukocytes to migrate through the endothelium. The assay is designed with a 3 µm pore size cell culture insert, appropriate for most leukocyte migration. 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 evaluate the effects of various factors that influence the transendothelial process.
Precoated cell culture inserts are provided in the Millipore QCM Leukocyte Transendothelial Cell Migration Assay to significantly reduce assay time. Additionally, the assay allows quantitative analysis of leukocyte migration. Following incubation of leukocytes with the endothelial cell layer, migratory cells are stained and quantified with WST-1 reagent and measured using a spectrophotometer. Absorbance correlates with cell migration.
|Background Information||Leukocytes patrol the vascular system. They must migrate across endothelial barriers in order to recruit to sites of inflammation. This process involves a multistep cascade consisting of leukocyte rolling, adhesion, and transmigration. A quantitative assay for leukocyte transendothelial migration has been described using a modified Boyden chamber system (Roth et al. 1995, Ding et al. 2000). 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. Once a confluent layer of cells is established, leukocytes are then added onto the endothelial monolayer. Leukocyte migration across the endothelium is determined by measuring the number of cells that migrate between the endothelial cells, through the porous membrane, to the lower chamber.
|Materials Required but Not Delivered||1. Endothelial cells (for example: HUVECs, Cat No. SCCE001)
2. Endothelial cell culture medium (Cat. No. SCME001)
3. Leukocytes and cell culture medium
4. 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.
5. Serum-free medium, such as RPMI-1640, DMEM (Cat No. ES-101-B), MEM, etc. containing 0.5% BSA (Cat No. 82-046-4), Pen/Strep (Cat No. TMS-AB2-C).
6. Sterile PBS (Cat No. BSS-1006-B) to wash cells.
7. Distilled water (Cat No. TMS-006-B)
8. (Optional) Chemoattractant or pharmacological agent for addition to culture medium.
9. Low speed centrifuge and tubes for cell harvesting.
10. CO2 incubator appropriate for subject cells.
11. Microplate reader (420 - 480 nm detection) or a spectrophotometer.
|Safety Information according to GHS|
|Material Size||1 kit|
|Material Package||24 Assays|
QCM™ Leukocyte Transendothelial Migration Assay (Colorimetric, 24 Assays) SDS
|Reference overview||Pub Med ID|
|Regulation of chemokine-induced transendothelial migration of T lymphocytes by endothelial activation: differential effects on naive and memory T cells.|
Ding, Z, et al.
J. Leukoc. Biol., 67: 825-33 (2000) 2000
|Characterization of transendothelial chemotaxis of T lymphocytes.|
Roth, S J, et al.
J. Immunol. Methods, 188: 97-116 (1995) 1995
|QCM Leukocyte Transendothelial Migration Assay - Colorimetric|