I want to lyse PBMCs. in that case do i need to put 100 microlitre for 1 ml? Is there is a variation of sigma cocktail according to cell type?
Ok, if P8340 is the correct cocktail, then it is written there to use 1 mL for 100 mL (20 g liver weight) meaning it is a 100x stocking solution. This leads to the conclusion that you have to use 10 µL for 1 mL of lysis buffer in case you have less than 0.2 g (200 mg) organ weight. Since you are using cells I guess you are solving something like 10 µL pellet in roughly 50-100 µL lysis buffer. Therefore, a 1:100 dilution of the protease cocktail should be fine. In case your protein is higher concentrated you need to add more inhibitor. I hope this will help! Good luck!
Tnx for suggestions. Ya, its a mistake the correct catalog number is P8340.
I had read the sigma datasheet where it was wriitten "One ml of the cocktail solution is recommended for 10 ml of cell lysate obtained from CHO cells at a cell density of 100 million cells per ml". So the solution is 10x in that case. In case of liver tissue, the datasheet recommended 1ml for 100ml.
I am using 10 microlitre cocktail in 100 microlitre of cell lysate.
Since the cocktail I purchased comes as 1ml. So I was worried, if I will use 10 microlitre for 100 microlitre of lysate, it will get finished soon. I was just thinking am I using too much cocktail. 10 microlitre for 100 microlitre of lysate.
10 microlitre for 100 microlitre of lysate sounds for me like quite a lot specifically in relation to the fact that it is solved in DMSO, which is not a harmless substance. I think you can use less or switch over to another company which are making it more clear like Roche (50x solved in water when you order the tabs). Nevertheless, it still depends on how high concentrated your protein lysate is.
As this is 100x sol. Then, what should be diluting solution for this cocktail. Can we dissolve in PBS or some other diluting agent.
Diluting solution will be your buffer which ever lysate buffer you are using for your prep. If you are using Ripa buffer then add 10ul in each 1ml of your buffer.
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