When designing a cytoprofiling experiment using TetonTM or Teton AtlasTM using iPSC derived mesenchymal stem (MSC) or neural progenitor (NPC) cells, there are some considerations for choosing the best slide coating. Element Biosciences offers bare glass slide kits as well as Poly-L-Lysine coated slide kits that can be combined with custom coatings like Matrigel, Laminin, and/or Poly-D-Lysine.
MSCs and NPCs can have specific conditions for adequate cell growth prior to a cytoprofiling run. We have tested various conditions and cell coatings with MSCs and NPCs to ensure cytoprofiling run success.
Mesenchymal Stem Cells
Three different combinations of slide coatings were tested with bone marrow derived mesenchymal stem cells at various concentrations -- Matrigel on Element’s bare glass slide kit, Matrigel added to Element’s Poly-L-Lysine (PLL) slide kit, and Matrigel and 0.01% Poly-D-Lysine (PDL) on Element’s bare glass slide kit. Cells were all seeded at 5k cells/well. All surfaces and Matrigel concentrations were able to successfully grow mesenchymal stem cells.
Other coatings including Matrigel Growth Factor Reduced (GFR) on Element PLL, Matrigel Growth Factor Reduced (GFR) on Element’s glass slide kit, Laminin on Element’s glass slide kit, and Matrigel and Laminin plus 0.01% PDL on Element’s glass slide kit were also tested with similar efficacy. Please see the concentration table in the Summary of Recommendations section below for recommendations when using any of these coatings.
Neural Progenitor Cells
The same coatings and concentrations were tested using neural progenitor cells. Cells were all seeded at 10k cells/well. We found that Matrigel on bare glass led to cell loss during fixation, especially as Matrigel concentration increased. Matrigel on Element’s PLL slide kit and Matrigel with 0.01% PDL on Element’s bare glass slide kit did not observe the same cell loss during fixation, but did lead to some cell aggregation. Cell aggregation can lead to issues with segmentation and can negatively affect your cytoprofiling run as a result.
Additional surface options were tested with neural cell growth factors including Matrigel Growth Factor Reduced (GFR) on bare glass, Matrigel GFR on Element PLL, and Laminin on bare glass. Cells were seeded at 6k cells/well for 18 hours.
We identified that Matrigel GFR (Corning catalog number 354231) works very well with neuronal cells. Laminin is also a good alternative for neuronal cell growth.
Summary of Recommendations
Through this optimization, we’ve included a table of down selected surface coatings and working concentrations for NPCs and MSCs.
Surface Coating Type | MSCs: Working Concentration Range (mg/mL) | NPCs: Working Concentration Range (mg/mL) |
Element PLL + Matrigel | 0.05-1.0mg/ml | 0.25-1.0mg/ml |
Element PLL + Matrigel GFR | 0.05-1.0mg/ml | 0.05-1.0mg/ml |
Matrigel GFR | 0.05-1.0mg/ml | 0.05-1.0mg/ml |
Laminin | 0.05mg/ml | 0.05mg/ml |
Matrigel/Laminin on 0.01% PDL | 0.05-1.0mg/mL (Matrigel) | 0.5-1.0mg/mL (Matrigel) |
Matrigel | 0.05-1.0mg/ml | Not effective |
Geltrex | 0.25mg/ml | Not tested |
Matrigel Growth Factor Reduced is the optimal coating for NPCs. Any major cell loss during fixation suggests that cell coating, fixation protocol, and/or handling needs to be optimized further. We recommend optimizing working concentrations using our Teton Custom Antibody Screening Kit, Teton Onboard Cell Paint Imaging Kit or the Teton Cell Paint Probe Kit for best results.