We cultured cells on de-epitheliased amniotic membrane on the stromal side. However, when we want to isolate the cells, RNA concentration was low, although under light microscope and scanning electron microscope observation, we can see the cells.
We also believe the cells penetrated into the membrane through the gap of stromal surface.
Badreddine Mechri
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Laurence Stuart Dawkins-Hall
Dear Azlina
- as indicated by the papers above and from personal experience, yes there is
- In essence, unless you effectively remove thge ECM by enzymatic digestion, resulting RNA can have residual matrix polymeric impurities that can interfere with both RT and PCR
- Thus, on one occasion when harvesting cells from Hydrogels by standard washing procedures followed by standard RNA extraction methods, when I evluated primer efficincy for qPCR by serial dilution of cDNA derived from RNA harvested in this way the efficiencies were only about 60%: This is too low to derive meaningful Gene expression data
- Thus, purify RNA as above to render PCR amplification efficient; that is > 85%
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