CytoSelect™ 24-Well Cell Invasion Assay (Basement Membrane, Colorimetric Format)

Details for Product No. ABIN2344859, Supplier: Log in to see
Reactivity
Mammalian
Application
Cellular Assay (CA)
Options
Supplier
Log in to see
Supplier Product No.
Log in to see
Request

Get this product for free

It's quick and easy to submit your validation proposal. I want to validate this product

Learn more

Purpose The CytoSelect™ Cell Invasion Assay Kit contains polycarbonate membrane inserts (8 μm pore size) in a 24-well plate. The upper surface of the insert membrane is coated with a uniform layer of dried basement membrane matrix solution. This basement membrane layer serves as a barrier to discriminate invasive cells from non-invasive cells. Invasive cells are able to degrade the matrix proteins in the layer, and ultimately pass through the pores of the polycarbonate membrane. Finally, the cells are removed from the top of the membrane and the invaded cells are stained and quantified. 2
Brand CytoSelect™
Sample Type Serum, Cell Samples
Analytical Method Quantitative
Detection Method Colorimetric
Characteristics CytoSelect™ Cell Invasion Assay Kit utilizes basement membrane-coated inserts to assay the invasive properties of tumor cells. This Trial Size kit contains sufficient reagents for the evaluation of 4 samples.
Components
  1. ECM Invasion Chamber Plate : One 24-well plate containing 4 ECM-coated cell culture inserts.
  2. Cell Stain Solution : One 4 mL bottle
  3. Extraction Solution : One 4 mL bottle
  4. Cotton Swabs: 40 each
  5. Forceps: One each
Material not included
  1. Invasive cell lines
  2. Cell culture medium
  3. Serum free medium, such as DMEM containing 0.5 % BSA, 2 mM CaCl2 and 2 mM MgCl2
  4. Cell culture incubator (37 °C, 5 % CO2 atmosphere)
  5. Light microscope
  6. 96-well microtiter plate
  7. Microtiter plate reader
Background The ability of malignant tumor cells to invade normal surrounding tissue contributes in large part to the significant morbidity and mortality of cancers. Invasiveness requires several distinct cellular functions including adhesion, motility, detachment, and extracellular matrix proteolysis. Metastatic cells produce many proteolytic enzymes (e.g. lysosomal hydrolysates, collagenases, plasminogen activators) while the expression of certain cell surface protease receptors is also increased.
Application Notes Optimal working dilution should be determined by the investigator.
Comment

  • Fully quantify cell invasion with no manual cell counting
  • Plate inserts are precoated with ECM basement membrane

Plate Pre-coated
Assay Procedure
  1. Under sterile conditions, allow the invasion chamber plate to warm up at room temperature for 10 minutes.
  2. Rehydrate the basement membrane layer of the cell culture inserts by adding 300 μL of warm, serum-free media to the inner compartment. Incubate at room temperature for 1 hour.
  3. Prepare a cell suspension containing 0.5-1.0 x 106 cells/mL in serum free media. Agents that inhibit or stimulate cell invasion can be added directly to the cell suspension. Note: Overnight starvation may be performed prior to running the assay
  4. Carefully remove the rehydration medium (step 2) from the inserts without disturbing the basement membrane layer. Note: It will not affect the assay performance if a small amount of rehydration medium is left in the compartment
  5. Add 500 μL of media containing 10 % fetal bovine serum or desired chemoattractant(s) to the lower well of the invasion plate.
  6. Add 300 μL of the cell suspension solution to the inside of each insert.
  7. Incubate for 24-48 hours at 37 °C in 5 % CO2 atmosphere.
  8. Carefully aspirate the media from the inside of the insert. Wet the ends of 2-3 cotton-tipped swabs with water, flatten the ends of the swabs by pressing them against a clean hard surface, and gently swab the interior of the inserts to remove non-invasive cells. Take care not to puncture the polycarbonate membrane. Be sure to remove cells on the inside perimeter of the insert.
  9. Transfer the insert to a clean well containing 400 μL of Cell Stain Solution and incubate for 10 minutes at room temperature.
  10. Gently wash the stained inserts several times in a beaker of water. Allow the inserts to air dry.
  11. (optional) Count invasive cells with a light microscope under high magnification objective, with at least three individual fields per insert.
  12. Transfer each insert to an empty well, adding 200 μL of Extraction Solution per well, then incubating 10 minutes on an orbital shaker.
  13. Transfer 100 μL from each sample to a 96-well microtiter plate and measure the OD 560nm in a plate reader. 4
Restrictions For Research Use only
Storage 4 °C
Storage Comment Store all components at 4°C.
Product cited in: Rodríguez-Mateo, Torres, Gutiérrez, Pintor-Toro: "Downregulation of Lnc-Spry1 mediates TGF-β-induced epithelial-mesenchymal transition by transcriptional and posttranscriptional regulatory mechanisms." in: Cell death and differentiation, Vol. 24, Issue 5, pp. 785-797, 2017 (PubMed).

Huang, Luo, Han, Huang, Tang, Wu: "MiR-223/PAX6 Axis Regulates Glioblastoma Stem Cell Proliferation and the Chemo Resistance to TMZ via Regulating PI3K/Akt Pathway." in: Journal of cellular biochemistry, 2017 (PubMed).

Ma, Yu: "Over-Expression of TBL1XR1 Indicates Poor Prognosis of Serous Epithelial Ovarian Cancer." in: The Tohoku journal of experimental medicine, Vol. 241, Issue 3, pp. 239-247, 2017 (PubMed).

Oba, Nakahara, Hashimoto-Hachiya, Liu, Abe, Hagihara, Yokomizo, Furue: "CD10-Equipped Melanoma Cells Acquire Highly Potent Tumorigenic Activity: A Plausible Explanation of Their Significance for a Poor Prognosis." in: PLoS ONE, Vol. 11, Issue 2, pp. e0149285, 2016 (PubMed).

Slusser-Nore, Larson-Casey, Zhang, Zhou, Somji, Garrett, Sens, Dunlevy: "SPARC Expression Is Selectively Suppressed in Tumor Initiating Urospheres Isolated from As+3- and Cd+2-Transformed Human Urothelial Cells (UROtsa) Stably Transfected with SPARC." in: PLoS ONE, Vol. 11, Issue 1, pp. e0147362, 2016 (PubMed).

Tansi, Rüger, Böhm, Kontermann, Teichgraeber, Fahr, Hilger: "Potential of activatable FAP-targeting immunoliposomes in intraoperative imaging of spontaneous metastases." in: Biomaterials, Vol. 88, pp. 70-82, 2016 (PubMed).

Nam, Kim, Park, Bang, Jin, Lee, Kim, Han, Im, Kim, Oh, Bang: "Src as a Therapeutic Target in Biliary Tract Cancer." in: Molecular cancer therapeutics, Vol. 15, Issue 7, pp. 1515-24, 2016 (PubMed).

Calabriso, Massaro, Scoditti, Pellegrino, Ingrosso, Giovinazzo, Carluccio: "Red Grape Skin Polyphenols Blunt Matrix Metalloproteinase-2 and -9 Activity and Expression in Cell Models of Vascular Inflammation: Protective Role in Degenerative and Inflammatory Diseases." in: Molecules (Basel, Switzerland), Vol. 21, Issue 9, 2016 (PubMed).

Ohnishi, Iwatsuki, Ishihara, Ohkawa, Kinoshita, Shinzawa, Fujimoto, Yoshimine: "Promotion of astrocytoma cell invasion by micro RNA-22 targeting of tissue inhibitor of matrix metalloproteinase-2." in: Journal of neurosurgery. Spine, Vol. 26, Issue 3, pp. 396-403, 2016 (PubMed).

Wei, Sun, Li: "MicroRNA-215 enhances invasion and migration by targeting retinoblastoma tumor suppressor gene 1 in high-grade glioma." in: Biotechnology letters, Vol. 39, Issue 2, pp. 197-205, 2016 (PubMed).

Devis, Moiola, Masia, Martinez-Garcia, Santacana, Stirbat, Brochard-Wyart, García, Alameda, Cabrera, Palacios, Moreno-Bueno, Abal, Thomas, Dufour, Matias-Guiu, Santamaria, Reventos, Gil-Moreno, Colas: "Activated leukocyte cell adhesion molecule (ALCAM) is a marker of recurrence and promotes cell migration, invasion, and metastasis in early-stage endometrioid endometrial cancer." in: The Journal of pathology, Vol. 241, Issue 4, pp. 475-487, 2016 (PubMed).

Chen, Xu, Nagati, Hogg, Das, Gerard, Garcia: "The Acetate/ACSS2 Switch Regulates HIF-2 Stress Signaling in the Tumor Cell Microenvironment." in: PLoS ONE, Vol. 10, Issue 2, pp. e0116515, 2015 (PubMed).

Modali, Parekh, Kebebew, Agarwal: "Epigenetic regulation of the lncRNA MEG3 and its target c-MET in pancreatic neuroendocrine tumors." in: Molecular endocrinology (Baltimore, Md.), Vol. 29, Issue 2, pp. 224-37, 2015 (PubMed).

Hirata, Hinoda, Shahryari, Deng, Nakajima, Tabatabai, Ishii, Dahiya: "Long Noncoding RNA MALAT1 Promotes Aggressive Renal Cell Carcinoma through Ezh2 and Interacts with miR-205." in: Cancer research, Vol. 75, Issue 7, pp. 1322-31, 2015 (PubMed).

Li, Wu, Abbatiello, Wu, Kim, Sarkissyan, Sarkissyan, Chung, Elshimali, Vadgama: "Slug contributes to cancer progression by direct regulation of ER? signaling pathway." in: International journal of oncology, Vol. 46, Issue 4, pp. 1461-72, 2015 (PubMed).

Wang, Li, Bao, Wu, Du, Su, Zhang, Liang: "MicroRNA-873 (miRNA-873) inhibits glioblastoma tumorigenesis and metastasis by suppressing the expression of IGF2BP1." in: The Journal of biological chemistry, Vol. 290, Issue 14, pp. 8938-48, 2015 (PubMed).

Kang, Kang, Kim, Lee, Ha, Yeo, Kang, Moon, Lee: "Curcumin sensitizes human lung cancer cells to apoptosis and metastasis synergistically combined with carboplatin." in: Experimental biology and medicine (Maywood, N.J.), Vol. 240, Issue 11, pp. 1416-25, 2015 (PubMed).

Cheng, Zheng, Bu, Wu, Zhang, Xing, Wang, Hu, Du, Li, Li, Zhou, Wen, Ji: "LAPTM4B-35, a cancer-related gene, is associated with poor prognosis in TNM stages I-III gastric cancer patients." in: PLoS ONE, Vol. 10, Issue 4, pp. e0121559, 2015 (PubMed).

Desai, Kharade, Parekh, Iyer, Agarwal: "Pro-oncogenic Roles of HLXB9 Protein in Insulinoma Cells through Interaction with Nono Protein and Down-regulation of the c-Met Inhibitor Cblb (Casitas B-lineage Lymphoma b)." in: The Journal of biological chemistry, Vol. 290, Issue 42, pp. 25595-608, 2015 (PubMed).

Osawa, Yokoyama, Shigeto, Futagami, Mizunuma: "Decreased expression of carbonyl reductase 1 promotes ovarian cancer growth and proliferation." in: International journal of oncology, Vol. 46, Issue 3, pp. 1252-8, 2015 (PubMed).