= Bioassay
A \b[bioassay] measures a substance's biological activity through a specified biological response, such as a cellular response or an enzyme-catalyzed reaction. Its result depends on the test system and the conditions under which the response is observed. Chemical amount and biological activity are related questions, but measuring one does not automatically establish the other. The United States Pharmacopeia describes biological assays as important for determining potency and assuring activity of biological products.<Analysis of Biological Assays by United States Pharmacopeia>[Chapter 1034, Section 1, Introduction]{ref}
A quantitative bioassay specifies the preparation, applied dose or concentration, biological system, observation interval and readout. Comparing an unknown preparation with a reference in the same system can estimate <relative biological potency>. A <dose-response relationship> is more informative than a single treated-versus-untreated observation because it tests how response changes across the dose range.
For an original ideal model, let reference response at concentration $x$ be
$$
y_R(x)=\frac{x}{K+x},\qquad y_T(x)=\frac{\rho x}{K+\rho x}.
$$
Here $K$ is the reference half-response concentration and $\rho$ is the test preparation's relative potency under a common-shape model. At any chosen response strictly between zero and one, the equal-effect concentrations satisfy
$$
\frac{x_R}{x_T}=\rho.
$$
If the reference requires $10$ concentration units and the test requires $5$ for the same response, the modeled relative potency is $2$. This is an effect-based comparison, not evidence that the test contains twice the chemical mass.
<Bioassay parallelism> concerns whether the two response curves can be represented by a common shape shifted along log concentration. If slopes or attainable responses differ, an equal-effect concentration ratio can depend on the response level. Reporting one universal potency ratio would then hide a model failure. Reference preparations, biological replicates, blanks and procedural controls test different parts of the measurement.
An <MTT assay> supplies a concrete experimental example. The original method used tetrazolium reduction and a multiwell scanning <spectrophotometer>, with response depending on cellular activation.<Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays by T. Mosmann>[Abstract]{ref} Later work demonstrated colored <formazan> production without cells for selected test compounds.<Interference of plant extracts, phytoestrogens and antioxidants with the MTT tetrazolium assay by Regina Bruggisser and others>[Abstract, cell-free interference tests]{ref} The optical readout therefore needs a response model and appropriate controls before it is interpreted as <cell viability>. A reproducible color measurement is not by itself validation of the intended biological conclusion.
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