Please can you tell me the ratio of that substance.
Depends on what you are trying to accomplish.
For solving the problem of getting cells to stick to plastic, you could try fibronectin or vitronectin (they enhance adhesion and spreading) or poly l-lysine (cells adhere but don't spread well).
If your question is cell behavior on plastic versus an extracellular matrix (ECM), Matrigel would be a good, but expensive, choice, as it is essentially a solubilized (ECM) with all sorts of ECM proteins in a physiological ratio. But what collagen are you writing about? Collagen I (interstitial) may be less appropriate for your question than Collagen IV (basement membrane).
I agree with the previous comment. You can coat with almost anything (and certainly can coat with any ECM protein). However, what you coat with will drive the phenotype of the cells that adhere. Poly-L-lysine gives you a generic coating that things will stick to, but in a non-physiologic way. Integrins aren't activated and classical cell-ECM interactions are absent. Matrigel is expensive and "physiologic" but also may have growth factors like TGFR-beta and has a mix of different ECM proteins that may (or may not) resemble the physiologic state of the cells you are interested in. You can also coat with purified ECM proteins including various collagens, laminin, fibronectin (plasma or tissue), fibrinogen depending on what you're interested in. You will find important differences in phenotype and cell signaling depending on what you choose. For coating plastic dishes, ideally use bacteriologic plastic rather than tissue culture plastic. For many years, we've successfully used an ELISA-type bicarb-based coating buffer with great success that I learned from Joe Madri. (See Human enterocyte (Caco-2) migration is modulated in vitro by extracellular matrix composition and epidermal growth factor. Basson MD, Modlin IM, Madri JA. J Clin Invest. 1992 Jul;90(1):15-23.) Although originally described on bacteriologic plastic, we have successfully used the technique it for various other membranes as well.
I agree with all answers above.. it depends what type of cells you are culturing.
For example, I culture stem cells using CellStart as a coating matrix in xeno free cultures instead of Matrigel. It's expensive but works for me.
I hope that helps.
cheers
Thank you for answer Im going to isolate rat liver primary cells
I am not an expert on primary hepatocyte culture, unfortunately:-) However, I think there are a number of protocols with isolation tips and ones with culture conditions using rat tail collagen as the matrix. Might also be commercial kits available using 3D systems if you need the most differentiated cells possible.
For a gelled substrate?
For every ml of gel, I used to mix together 0.1 ml sterile 1N NaOH [to raise pH], 0.1 ml 10x PBS [to raise salt concentration] + 0.8 ml commercial 2.5-3 mg/ml rat tail collagen ON ICE. I would pipet 1 ml/35 mm dish, swirl to coat the bottom, and gel @ 37ºC for ~30 min. I would then equilibrate the gel with growth media for ~30 min before seeding cells. If you use 10x DMEM instead of 10x PBS, your could probably skip the equilbration step.
I would think so. The cells might not spread well though
Here is a reference for culturing rat hepatocytes on different types of collagen substrates (with pictures). Hope it helps:-)
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