Optimizing Matrigel Surface Coatings for Teton CytoProfiling Runs

When designing a cytoprofiling experiment using Teton or Teton AtlasTM, a key component is selecting the best slide coating for your cell type. Element Biosciences provides a slide kit that is pre-coated with Poly-L-Lysine, or, alternatively, an uncoated slide kit that can be used with various custom coatings including Matrigel.

The most ideal Matrigel coating is dependent on the cell type. To optimize your Matrigel concentration and seeding conditions, we recommend leveraging the Teton Custom Antibody Screening Kit, Teton Onboard Cell Paint Imaging Kit or the Teton Cell Paint Probe Kit. These kits allow you to visualize the cell membrane and nuclear labels under fluorescence to better assess performance and workflow prior to preparing cells for a full Teton or Teton Atlas run.

Test a range of concentrations diluted in chilled 1x PBS. We generally see success within a range of 0.05 – 1mg/ml although some cell lines may require a higher concentration. Overall, the concentration of Matrigel should be <2.5mg/ml to retain an even coating and reduce background staining. Additionally, some cell types may benefit from a dual coating of Matrigel on top of a Poly-L-Lysine pre-coated slide. An example setup in a 12-well slide kit is shown below to test a range of concentrations in replicates:

1

2

A

0.05 mg/ml

1.00 mg/ml

B

0.10 mg/ml

0.75 mg/ml

C

0.25 mg/ml

0.50 mg/ml

D

0.50 mg/ml

0.25 mg/ml

E

0.75 mg/ml

0.10 mg/ml

F

1.00 mg/ml

0.05 mg/ml

Ensure the Matrigel remains cold when preparing dilutions and applying to the slide surface – if Matrigel is too warm, clumps will form across the well. We recommend using Matrigel Basement Membrane Matrix, Phenol Red-free (Corning, 356237). Different Matrigel products could affect the coating. If testing a Matrigel product outside of our recommendation, set up a control and ensure that the Matrigel is Phenol Red-free as Phenol Red could affect imaging.

Capture brightfield images across a range of concentrations before and after fixation. Visually inspect the images to ensure that the coated surface maintains a consistent monolayer free of peeling or cracking. Confirm there is minimal background signal from the Cell Paint Probes. The cells should be well adhered to the surface with minimal cell loss. Compare confluency and concentration - you may notice higher confluency with lower Matrigel concentrations.

If there is cell loss prior to fixation, then likely the cells are not compatible with the surface. Alternatively, if cell loss is seen after fixation, then we can look to cell handling and/or weak cellular attachment to the surface.

1: Example of peeling and high background signal due to high Matrigel concentration
2: Zoomed in example of good cell adherence, confluency and minimal background.

For more information, reach out to your Field Applications Scientist or Element Biosciences Support at support@elembio.com.

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