I want to assay cell viability with MTT powder from Merck company. What is best solvent of MTT powder for cell viability assay?
Thanks.
We dissolve 1G MTT (Sigma, M5655-1G) in 200 ml PBS (5 mg/ml). Aliquot it in 5 ml tubes and store it at -20. Add 20 ul to 200 ul media containing tested cells plus medium (after incubation time). Then add 50 ul MTT stop solution (formazon solvent). Incubate for 4 hours at +37. Read OD at 570 nM.
I agree, in PBS. You can make a solution at 5 mg/ml. After vortex it, sonication allows complete solubilization.
Inject 100 µl of this MTT stock solution on cell (for 1 ml medium culture).
Sincerly
PBS (pH 7.4) is the best solvent. The powder should be dissolved in a dark colored bottle or aluminum foil wrapped falcon tube. A concentration at 5.5 mg/ml gives better result. The best brand is from Sigma Aldrich. TQ
MTT powder should be prepared in a physiologically balanced solution, typically PBS.
I use it at a concentration of 0.5mg/ml
you can either use PBS or even DMEM without phenol red (transperent solution)... you can use 10% of 5mg/ml MTT (0.5mg/ml) and add 100 micro litre in 96 well plate after discarding the medium from wells...filter MTT solution prior use (0.2 micron). keep it for 4 hours incubation and discard it.
to dissolve MTT (formazan cryastals) add either acidified isopropyl alcohol or DMSO.
mix it properly and read it at 550 nm
Hi all of you.
Is absolutely necessary to filter MTT with the solution in which was dissolved, either PBS or culture media?
Thanks a lot!
I dissolve in PBS at 0.5mg/ml final concentration and filter MTT solution using 0.22um filter.
it mix at slow speed on a magnetic stir plate for 15 minutes until the MTT powder is completely dissolved.
I use PBS to make MTT solution at a stock concentration of 5mg/mL and use syringe filter not just to sterilize it, but to remove any insoluble particulates. I use it by diluting the stock 1/5 in media.
I usually use 5mg/ml concentration of MTT.
Would the concentration of MTT be a major factor? If so, then how?
MTT Stock solution 5 mg/ml of PBS and use 10 ul /well of 96 well culture plat(100ul) cell culture.
We dissolve 1G MTT (Sigma, M5655-1G) in 200 ml PBS (5 mg/ml). Aliquot it in 5 ml tubes and store it at -20. Add 20 ul to 200 ul media containing tested cells plus medium (after incubation time). Then add 50 ul MTT stop solution (formazon solvent). Incubate for 4 hours at +37. Read OD at 570 nM.
Filtration of MTT solution after preparation is very important, however it depends on the purity of MTT solution. For example if you dissolve MTT from Sigma with cat. Number:M2128, you should filter it.
But the same product with cat. Number from sigma: M5655, as it is mentioned suitable for cell culture, it does not need filteration.
I dissolved my mtt powder in isopropanol and got strange results (my controls which had only medium, showed less color than my conditions). I am not sure these results is because of Iso-propanol or something went wrong to my cells. Please tell me your suggestions
Hi,
Usually we using PBS to dissolve MTT powder. The concentration should be from 1 to 5 mg/ml.
In my lab, we dissolve MTT in PBS (5mg/ml). Remember to vortex vigirously, then use the sterile filter (0,5µm) to remove the remained crystals and sterile it.
I use 5 mg/mL concentration of MTT by disolving 500 mg in PBS (100 mL). Once filtered, I store it at 4 ºC.
Hi Marwa,
PBS (Phosphate Buffer Saline 1X) of pH 7.4 is the best solvent for MTT.
Regards,
Arun AB
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