How does ‘Easy Coat’ work?
Easy Coat hydrogels are populated with quinone groups, which form covalent bonds with molecules containing a primary amine, thiol, or strong nucleophile – essentially any protein. Be aware that once all the binding sites are reacted, additional molecules will not bind directly to the hydrogel.
Can I detach my cells with trypsin?
Yes, but prior to adding trypsin, incubate the hydrogel in serum-free media or buffer for ≥30 min at 37°C to remove serum absorbed in the gel. To bypass this step, use TrypLE™ (Life Technologies), which dissociates cells in serum-containing conditions.
How do I isolate RNA?
We recommend using a spin-based kit such as RNeasy© (Qiagen). Incubate the hydrogel with lysis buffer for 10 min on ice, collect the lysate, and repeat to recover RNA absorbed in the gel.
Can I scrape my cells off the hydrogel?
A cell scraper with a silicone rubber blade will do the trick (available from CytoOne©). Scrape with gentle, short strokes to avoid disrupting the hydrogel.
Can I fix my cells?
Softwell is compatible with fixatives such as formalin, glutaraldehyde, and methanol. Note that dehydrating agents will cause the gel to turn opaque, but this is reversible.
Can I image my cells?
The hydrogels are transparent and compatible with brightfield, phase contrast, and fluorescence microscopy. The combined thickness of the hydrogel and substrate is within the focal range of 40X objectives. For higher magnifications, use Softslip hydrogels, which can be inverted onto a coverslip.
Is Softwell compatible with my assay?
The hydrogels are thin, transparent, and bound to the well, which maximizes compatibility with a wide range of cell-based assays. To ensure removal of unbound detection molecules, perform wash steps 3x longer than directed by standard protocols.
Is Softwell like Matrigel™?
Hydrogels <1 kPa are as soft as Matrigel. But whereas Matrigel is comprised of tumor-derived proteins and growth factors, Softwell is based on a synthetic polymer.