Matrigengel Matrix-0827045-Mogengel

Description

Basement membranes are continuous sheets of specialized extracellular matrix that form an interface between endothelial, epithelial, muscle, or neuronal cells and their adjacent stroma. Basement membranes are degraded and regenerated during development and wound healing. They not only support cells and cell layers, but they also play an essential role in tissue organization that affects cell adhesion, migration, proliferation, and differentiation. Basement membranes provide major barriers to invasion by metastatic tumor cells.

Mogengel Matrigengel Matrix is a soluble form of basement membrane purified from Engelbreth-Holm-Swarm (EHS) tumor.Mogengel Matrigengel Matrixat 37 °C to form a reconstituted basement membrane. The major components of Mogengel Matrigengel Matrix  include laminin, collagen IV, entactin, and heparin sulfate proteoglycan.

Intended Use

2D or 3D cultures related to cell proliferation or differentiation,as well as study of cell morphological,Commonly used experiments are on cell invasion , angiogenesis and organoid culture, among others.

Materials Provided

Product Parameter

Source: Mouse Tumor

Appearance：

① Color: phenol-containing red matrigengel is yellow-pink, and phenol-free red matrix gum is translucent light yellow;

② Form: standard matrigengel dissolved at 4℃, a transparent liquid state; High concentration matrigengel after 0℃ solution, transparent liquid state, 4℃ for a long time to show a semi-gel.

Concentration: Protein concentration ranges from 8 to 26mg/mL

Endotoxin:≤ 4.5EU/ mL

Gel time: 5-30min gel at room temperature, the speed of gel formation is accelerated when the temperature is from 22°C to 37°C.

Material Qualifications

l Routine screening of mouse colony pathogensby mouse antibody product (MAP) tests

l Testing for bacteria, fungi and mycoplasmato ensure negative results

l Extensive PCR testing for a variety of pathogens including LDEV to ensure strict control of raw materials used in the production process

l Extraction from LDEV-free mouse tumor cells

l Gel stability testing at 37℃ for 14 days

l Detection of endotoxin levels using serological methods

l Biological function verification ofeach lot (organoid culture and differentiation experiments; Subcutaneous tumor formation test; Stem cell culture; Angiogenesis experiment etc.)

Coating Procedures

Thaw Matrigengel Matrix overnight at 2-8 °C. Refrigerator temperatures may vary, therefore it is recommended to keep Matrigengel on ice in a refrigerator during the thawing process. Thawed Matrigengel solidifies quickly at temperatures above 15 °C; when working with Matrigengel,

keep it on ice to prevent untimely gelling.

There are many applications for Matrigengel which require different thicknesses and concentrations. A thick gel is needed for applications such as endothelial cell formation of capillary-like structures (Tube Formation Assay), the differentiation of rat aorta tissue into capillary-like structures (Aortic Ring Assay), epithelial organoid formation, or tumor organoid formation. Some applications, such as propagation of primary cells, require a thin layer coating and not a thick gel; therefore, the thin layer method should be used.

Thick Gel Method:

1. Thaw Matrigengel as stated above.

2. Mix Matrigengel by slowly pipetting solution up and down; be careful not to introduce air bubbles.

3. Pipette 200-300 μL per cm² onto the growth surface.

4. Place coated object at 37 °C for 30 minutes.

5. Coated objects are ready for use.

Thin Layer Method (non-gelling):

1. Thaw Matrigengel as stated above.

2. Mix Matrigengel by slowly pipetting solution up and down; be careful not to introduce air bubbles.

3. Dilute Matrigengel to desired concentration in cold serum-free medium. A 1:100 dilution is recommended for the propagation of primary cells. Empirical determination of the optimal coating concentration for your application may be required.

4. Add a sufficient amount of solution to cover the entire growth surface area. A volume of 300 µL per cm² is recommended.

5. Incubate coated object at room temperature for one hour.

6. Aspirate coating solution and immediately plate cells. Do not allow coated surface to dry out.

Example for PROCEDURES

Angiogenesis assay

1. Equipment, reagents and consumables

1.1 Equipment: Biosafety cabinet, pipette, carbon dioxide incubator, inverted microscope, centrifuge(Low-speed)

1.2 Reagent: DMEM, ECM complete medium,Trypsin Solution, 1×PBS solution

1.3 Consumables: sterile pipette tips; 96-well cell culture plate; Sterile EP tube and other consumables. (Or be adjusted according to the experimental design).

2. Experimental contents and methods

2.1 Preparation before Experiment

2.1.1 Put the Matrigengel in the ice box and put it in the refrigerator at 4℃ so that the Matrigengel can slowly melt overnight; (Do not allow this product to warm up above 4°C during manipulation. Keep the product on ice and dilute using ice-cold solutions or cell suspensions.)

2.1.2 Consumables or reagents that come into contact with Matrigengel, such as sterile centrifuge tube, sterile pipette tips and DMEM , were be pre-cooled at 4℃ in advance;

2.2 Plate coating procedure

2.2.1 Prepare the EP tube for placing on ice. Add each component according to the following table.

Note:The Ratio 2:1 is appropriate in most case.

2.2.2 After mixing the above components in sequence, add 50µL/ well to the 96-well plate, ( two repeated wells are be recommended), mark the information and the experiment date, and solidified in a 37℃ incubator for at least 1 hour.

2.3 Incubate the HUVEC

2.3.1 HUVEC were selected for pancreatic enzyme digestion. Cell density was adjusted to 4×10⁵cells /mL by using ECM medium.

2.3.2 Take the “2.2.2” Coated 96-well plate and add the cell suspension in a volume of 50 μL per well（It means that the final number of HUVEC is 2×10⁴）.

2.3.3 The results of vascular structures can be observed after after 4 h and 24 h incubation in a carbon dioxide incubator at 37 °C.