Millicoat™ Human Collagen Type IV Coated Strips (96-Wells)

Millicoat™ Human Collagen Type IV Coated Strips (96-Wells)

Chemicon (Millipore)

1 plate

Cell adhesion plays a major role in cellular communication and regulation, and is of fundamental importance in the development and maintenance of tissues. Scientists are continually examining the adhesion and migration of many diverse cell types on various extracellular matrix (ECM) component proteins. Millicoat™ Cell Adhesion Strips are provided as 12 x 8-well removable strips in a plate frame for convenience and flexibility in designing assays. The wells in rows A - G have been coated with Human Collagen Type IV. Row H of each strip is coated with BSA which serves as a negative assay control. Cells are seeded onto the coated substrate. Subsequently, adherent cells are fixed and stained. Relative attachment is determined using absorbance readings.

Enzyme Assay

PROCEDURE:

NOTE: Optimal assay timing and performance may vary for different cell lines but generally can be obtained using subconfluent cell cultures in the assay described below. Subconfluent cultures can be achieved by splitting cells 1 to 2 days prior to performing the assay.

1. Rehydrate the strips with 200 mL of PBS per well for at least 15 minutes at room temperature. Remove the PBS from the rehydrated strips.

2. Prepare a single cell suspension, preferably using a non-enzymatic dissociation buffer. Optimum cell density may be determined by titration of the cells. A common starting range is between 1x10E05 to 1x10E07 cells/mL.

3. Add 100 mL of the diluted cell suspension to each well. Incubate the plate at 37°C for 45 minutes in a CO2 incubator. Gently wash the plate 2-3 times with PBS containing Ca2+/Mg2+ (200 mL/well).

4. Add 100 mL of 0.2% crystal violet in 10% ethanol to each well. Incubate for 5 minutes at room temperature. Remove the stain from the wells. Gently wash the strips 3-5 times with PBS (300 mL/well) to remove the excess stain.

5. Add 100 mL of Solubilization Buffer (A 50/50 mixture of 0.1M NaH2PO4, pH 4.5 and 50% ethanol) to each well. Allow strips to incubate and gently shake at room temperature until the cell-bound stain is completely solubilized; approximately 5 minutes.

6. Determine the absorbance at 540 - 570 nm on a microplate reader.

Human

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.

This gene encodes the major type IV alpha collagen chain of basement membranes. Like the other members of the type IV collagen gene family, this gene is organized in a head-to-head conformation with another type IV collagen gene so that each gene pair shares a common promoter.

FUNCTION: SwissProt: P02462 # Type IV collagen is the major structural component of glomerular basement membranes (GBM), forming a 'chicken-wire' meshwork together with laminins, proteoglycans and entactin/nidogen. Potently inhibits endothelial cell proliferation and angiogenesis. Inhibits angiogenesis potentially via mechanisms involving cell surface proteoglycans and the alpha and beta integrins of endothelial cells.
SIZE: 1669 amino acids; 160615 Da
SUBUNIT: There are six type IV collagen isoforms, alpha 1(IV)- alpha 6(IV), each of which can form a triple helix structure with 2 other chains to generate type IV collagen network.
SUBCELLULAR LOCATION: Secreted, extracellular space, extracellular matrix, basement membrane.
TISSUE SPECIFICITY: Highly expressed in placenta.
DOMAIN: SwissProt: P02462 Alpha chains of type IV collagen have a non-collagenous domain (NC1) at their C-terminus, frequent interruptions of the G- X-Y repeats in the long central triple-helical domain (which may cause flexibility in the triple helix), and a short N-terminal triple-helical 7S domain.
PTM: Lysines at the third position of the tripeptide repeating unit (G-X-Y) are hydroxylated in all cases and bind carbohydrates. & Prolines at the third position of the tripeptide repeating unit (G-X-Y) are hydroxylated in some or all of the chains. & Type IV collagens contain numerous cysteine residues which are involved in inter- and intramolecular disulfide bonding. 12 of these, located in the NC1 domain, are conserved in all known type IV collagens. & The trimeric structure of the NC1 domains may be stabilized by covalent bonds between Lys and Met residues.
DISEASE: SwissProt: P02462 # Defects in COL4A1 are a cause of brain small vessel disease with hemorrhage [MIM:607595]. Brain small vessel diseases underlie 20 to 30 percent of ischemic strokes and a larger proportion of intracerebral hemorrhages. Inheritance is autosomal dominant. & Defects in COL4A1 are a cause of porencephaly type 1 [MIM:175780]; also known as encephaloclastic porencephaly. Porencephaly is a term used for any cavitation or cerebrospinal fluid-filled cyst in the brain. Porencephaly type 1 is usually unilateral and results from focal destructive lesions such as fetal vascular occlusion or birth trauma. Inheritance is autosomal dominant.
SIMILARITY: Belongs to the type IV collagen family. & Contains 2 COL4C (collagen IV C-terminal) domains.

• Human Embryonic Stem Cells
• Mouse Embryonic Stem Cells
• Mesenchymal Stem Cells
• Neural Stem Cells
• Hematopoietic Stem Cells
• Epithelial Cells
• Pancreatic Stem Cells
• Induced Pluripotent Stem Cells

Maintenance & Expansion

Chemicon

P02462

Strips may be stored at 2-8°C in the foil pouch for at least 3 months. Unused strips may be placed back in the pouch for storage. Ensure that the desiccant remains in the pouch, and that the pouch is securely closed.

Chromogenic

96 wells

• COL4A1
• Arresten
• arresten

Millicoat™ Human Collagen Type IV Coated Strips (96-Wells)

NM_001845.4

Formerly under the CytoMatrix^TM brand name.