Bradford reagent could i use it after preparation or must incubate...

Mahmoud, the recommendation for incubation overnight is to allow complete dissolution of your dry reagents. If you are confident that your reagents are all thoroughly dissolved, you can use it immediately. Most formulations of a Bradford's reagent use a protein/reagent ratio of 1:25 (10uL/250uL; 100uL/2.5mL). Hope this helps, Mike