Currently, We are using free floating section in 12-well plates and small baskets to transfer tissues from one well to another. I have a few specific queries:
Could you please provide insights on the process you follow to clean the wells and baskets upon completing the procedure?
In terms of the baskets, do you maintain a consistent set throughout the entire procedure, or do you opt to replace them post-primary antibody incubation to prevent any antibody attachment to the basket mesh or walls?
Is it common practice for you to reuse plates for subsequent experiments?
I've been encountering challenges with the DAB staining phase. During the initial 5 minutes of DAB (with Ni) incubation, the sections appear to take on a light brownish hue, which progressively transitions to a bluish tone and finally settles into a dark blue shade after around 15 minutes.
1.I usually clean the baskets in PBS+detergent and rince in TBS. Wash them. in a beaker with a magnetic agitator and change the solution frequently.
2.You can keep the baskets throughout the procedure
3.You can reuse the plates after strong washes as in 1.
4.15 min is way too long as the reaction plateaus after 10min. I prefer to mount my tissue during the reaction under a binocular (protected from the DAB solution with plastic wrap). It will help you to stop the reaction when you see the positive signal.