Invasion Assay

Cell invasion is an important assay to study the cell mobility. The hydrogel matrix not only provide the structure scaffold but also induce cell-matrix interaction for cell mobility. VitroGel is a powerful system to study cell invasion. The tunable and functional hydrogel system can be manipulated to study many different factors such as mechanical strength, degradation, functional ligands and serum/growth factors/cytokine/chemokine that affect the cell mobility.  The cell invasion on VitroGel can be performed by using an insert or well plate for vertical movement, 3D hydrogel migration/invasion for horizontal movement, or 3D cell spheroid invasion assay.

Mechanical Strength
The cell mobility under different hydrogel mechanical strength can be studied by adjusting the hydrogel concentrations with VitroGel Dilution Solution. The typical gel strength range from 10 to 4,000 pa and the customized higher concentration hydrogel can get over 20,000 Pa.

Degradation
Using the VitroGel MMP, the matrix metalloproteinases sensitive hydrogel, the effect of the biodegradation on cell mobility can be studied. VitroGel MMP can also mix with other functional VitroGel system for a more complex microenvironment.

Functional Ligands
Multiple biological functional ligands are incorporated for different versions of VitroGel, which promote cell attachment, cell-matrix interactions and cell mobility. Taking the advantages of “Mix and Match” capability of VitroGel, scientists can also build a multi-functional VitroGel with different ratios of different functional ligands.

Serum/Growth Factors/Cytokine/Chemokines
The different concentrations of serum, growth factors and chemokines can be directly mixed with VitroGel or add to the outside of the insert well to induce the cell movement.

There are several ways to use VitroGel for invasion assay:

Insert or Well Plate
Using plate insert or well plate is a typical way to observe cell invasion. Besides the differences in medium components inside or outside of the insert, the tunable and functional VitroGel system can be manipulated to study many different factors such as mechanical strength, binding ligands on cell mobility.

Effects of FBS Concentration

Invasion of B35 neuroblastoma cells on VItroGel system.  VitroGel RGD was used as 2D hydrogel coating on the surface as cell insert. B35 cells were add directly on top of the hydrogel. The hydrogel was prepared at 1:3 dilution with 2% FBS. The cells was harvested from 2D culture plate with 10% FBS and resuspend in FBS free medium before adding to the top of hydrogel. The medium outside of the insert well contained 20% FBS. The cell invasion was induced by the different concentrations of FBS in cover medium, hydrogel and the medium outside of the insert. The images were taken at day 7.

 

Effects of Binding Ligands for Cell-Matrix Interactions

Invasion of U-87 MG glioblastoma cells on different functional VitroGel system.  VitroGel 3D was used as negative control and the RGD modified hydrogel (VitroGel RGD) was used to induce cell invasion. The hydrogel was prepared at 1:5 dilutions without FBS. The cells was harvested from 2D culture plate with 10% FBS and resuspend in culture medium with 1% FBS before adding to the top of hydrogel. The medium outside of the insert well contained 1% FBS and 2% human platelet lysate. The images were taken at the bottom of the insert well after 48 hrs.

 

Effects of Chemokine

Invasion of U-87 MG glioblastoma cells on VitroGel system induced by chemokines (CXCL12).VitroGel RGD was used to induce cell invasion. The hydrogel was prepared at 1:5 dilutions without FBS. The cells was harvested from 2D culture plate with 10% FBS and resuspend in culture medium with 1% FBS before adding to the top of hydrogel. The medium outside of the insert well contained 1% FBS and 10 ng CXCL12. The images were taken at the bottom of the insert well and the bottom of the well plate at day 7.
 

3D Hydrogel Migration/Invasion
The 3D horizontal invasion can be studied by encapsulating cells in hydrogel matrix. Scientists can adjust the properties of the hydrogel in the same well and observe the cell movement within hydrogel matrix.

3D Invasion of U-87 MG Glioblastoma Cells in VitroGel system.  U-87 MG cells were encapsulated in VitroGel RGD with 2% FBS and seed to a cell culture plate. After gel stable (30-60 min), use a micropipette to create a 5-10 µl hole by sucking the cell/hydrogel mixture out. Fill the hole with VitroGel RGD with 20% of FBS without cells. Add the cover medium with 10% of FBS on top of the hydrogel. The different concentrations of FBS in the hydrogel matrix would induce the cell invasion/migration within the hydrogel matrix. The cells movement was observed under a microscope with Z-stack function. The projecting images were created with different z levels.