When substrate solution is added to the reaction product, the enzyme substrate in the substrate solution decomposes, allowing the colouring agent to develop a colour.
H2O2 was added to 0.03%, the colour allowed to develop for 10 min and the slices washed extensively in dH2O.
Bound antibodies were visualised by incubation with DAB (liquid DAB cat K3468, Dako); the DAB was added to sections at 8 s intervals and the colour allowed to develop for exactly 3 min (ER β1) or 5 min (ER β2/ βcx).
The anthology for which he has provided his introduction is drawn from the workshop's writers, and the organisation is, he writes, "something right out of my wildest MFA dreams, where writers of colour could gather to develop our art in a safe, supportive environment.
Colour reagent, 3,3',5,5'-Tetramethylbenzidine (TMB) was dissolved in 0.05 M phosphate-citrate buffer, pH 5 containing 10% DMSO and added to each well, the colour was allowed to develop and the reaction stopped by adding H2SO4 to 0.4 M. The OD450 of each well was measured.
Subsequently, ceri-ammonium sulfate was added, and colour was allowed to develop at 250°C over 60 min.
The colour was allowed to develop at room temperature for 20 min before the detection of the absorbance value at 620 nm on the microplate spectrophotometer.
The colour was allowed to develop for 15 min at room temperature and the free phosphate levels were determined by measuring absorbance at 612 nm.
