B12 Reagent Explanation

One uncoated cup from Reagent 2 is taken
Serum added in it
Alkanine reagent Reagent 3-1 is added
Serum B12-Tc complex dissociate at alkaline pH and Tc get denatured (Reagent 3-1)
Reagent 3-2 (neutralizing reagent) added
On adding acid, pH is neutralized, but Tc remain denatured (Reagent 3-2)
Stabilizing reagent from Reagent-2 added to buffer at specific pH?????
Biotin-IF from Reagent 1 is added to uncoated cup
Sample B12 bind Biotin-IF. Some Biotin-IF remain unbound
[Sample B12-IF-Biotin complex]+[unbound Biotin-IF] from uncoated cup is taken and added to streptavidin-coated cup of Reagent-1
SampleB12-IF-Biotin complex bind well
B12-Free IF-Biotin bind well
B12-HRP comjugated added.
B12-HRP bind B12-Free IF-Biotin.
Excess B12-HRP washed
Reaction rate inversely proportion of B12 in serum