= Dissolved formazan recovery fraction
The \b[dissolved formazan recovery fraction] is the fraction of a specified amount of formed <formazan> that is represented as dissolved product in the solution being measured. Define
$$
r=\frac{n_{\mathrm{dissolved,read}}}{n_{\mathrm{formed}}},\qquad 0\le r\le1,
$$
for a model in which the same chemical product is conserved apart from incomplete dissolution or loss. The denominator and the stage at which it is established must be stated. Product formation, product recovery and the optical response per dissolved amount are distinct parts of an <MTT assay>.
The manufacturer's procedure forms product crystals before solubilization and multiwell optical measurement.<Cell Proliferation Kit I MTT instructions by Roche>[Section 3.1 and Figure 4, printed p. 9]{ref} A lower signal can therefore arise even when the same amount of product was initially formed.
For a homogeneous solution obeying the <Beer-Lambert law> in a fixed optical path,
$$
A-A_0=\epsilon\ell\frac{r n_{\mathrm{formed}}}{V}.
$$
If complete recovery would give net <absorbance> $0.40$, a matched observation of $0.30$ gives $r=0.75$ under that model. This inference requires the reference amount, <molar absorption coefficient>, volume and path length to be valid for the sample. A different solvent response or erroneous background can otherwise mimic incomplete recovery.
<Formazan recovery identifiability> is the central limitation: the optical observation alone determines the product $r n_{\mathrm{formed}}$, not its two factors. For example, $10$ arbitrary amount units formed with $r=0.5$ and $5$ units formed with $r=1$ give the same dissolved amount. Repeating the same optical measurement more precisely does not separate these explanations.
A known-product addition can test the recovery of that addition at a chosen processing stage. For matched optical conditions, the expected increment is $\Delta A_{\mathrm{ideal}}=\epsilon\ell\Delta n/V$, and the observed-to-expected increment estimates the recovery of the added product. An addition after dissolution does not test losses that occurred before it. Nor does a plateau after further mixing prove complete recovery without an independent amount reference.
An <MTT formazan solubilization check> should consequently record where product was added or formed, which liquid was retained, dilution, solvent and optical geometry. <Microplate fill-height optical cancellation> can change the volume dependence of the readout, but it does not restore product that was discarded or establish <cell viability>.
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