Hey, thank you for the detailed video. I have one question. When you add collagenase you mention it as 10mL of 60U/mL concentration But, when you are adding it to the small tissue pieces (present in the 50mL tube) you add two things using the pipette. Also, I understand you have prepared 60U/mL collagenase from the stock. Do you add it at that concentration directly to the cells or do you again dilute it while adding? Thank you!
@MedicinalChemistryCenter
Жыл бұрын
Dear Abhinaba Banerjee, We use ready-made collagenase solution in the laboratory (we use DMEM/F12 basal medium to dissolve the dry enzyme). The second reagent is calcium chloride, which is important for the work of collagenase. Good luck, MCC
@abhinababanerjee683
Жыл бұрын
@@MedicinalChemistryCenter Thank you for your reply! Also, from the stock to the working concentration do we dilute it in cell culture media? For the stock we use HBSS.
@helens6199
3 жыл бұрын
I noticed your bare wrists/arms and moving hands over the open samples - is there no risk of contamination in this kind of work? I'm a total beginner trying to understand the rules :)
@MedicinalChemistryCenter
3 жыл бұрын
There is no risk of contamination. Most importantly, wash your hands well up to the elbows before starting work.
@hariniamarnath2998
3 жыл бұрын
Woww tq for this step by step video.. Very good effort 👍👏👏
@donneestipona1720
4 жыл бұрын
This was a great, informative step-by-step video. Thank you for uploading. Quick question - is the interior of the culture flask coated with something to keep the cells adhered? It seems they don't come off until treatment with trypsin?
@MedicinalChemistryCenter
4 жыл бұрын
Hi Donne! Thanks for the comment. The flasks are not coated with the carrier. online-shop.eppendorf.ru/RU-ru/Kulturalnyj-plastik-110320/Rashodnye-materialy-dlja-raboty-s-kulturami-kletok-110321/Eppendorf-Cell-Culture-Flasks-PF-68138.html
@donneestipona1720
4 жыл бұрын
@@MedicinalChemistryCenter thank you for clarifying!
@somaiaassem2189
3 жыл бұрын
what is the function of cacl2 and collagenase
@MedicinalChemistryCenter
3 жыл бұрын
Thank you for your question. Collagenase is used to de-internationalize tissue and isolate tumor cells. Calcium ions are essential for collagenase to function.
@abhinababanerjee683
Жыл бұрын
As a second question, if you are trying to dissociate spheroids (of mouse astrocytes) embedded in collagen (2.4mg/ml) matrix, what working soln concentration would you recommend for collagenase? Also, would it be beneficial to use a cocktail of collagenase, trypsin, and acutase in this case?
@MedicinalChemistryCenter
Жыл бұрын
Unfortunately, we have no experience with astrocytes, but your cocktail of enzymes seems to me a tough option for the dissociation of spheroids. I think a mixture of collagenase and acutase will be enough. Good luck, MCC
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