= Western blot normalization
Total-<protein> <Western blot normalization> compares a target-band signal with a signal representing many transferred <proteins> in that lane. It is distinct from measuring one reference <protein>, and from measuring the target itself. Its denominator is a loading proxy: normalization estimates target abundance relative to detected total <protein>, rather than target amount per <biological cell>. Different sample compositions, selective <Western blot transfer> losses or unequal stain responses can still matter.
For an original calculation, suppose background-corrected target signals $I_A=12$, $I_B=24$ and total-<protein> signals $P_A=30$, $P_B=60$ are in their respective proportional response ranges. Then
$$
\frac{I_B/P_B}{I_A/P_A}=\frac{24/60}{12/30}=1.
$$
The invented signals double together, so this calculation supplies no evidence for a higher target fraction. With $I_B=24$ but $P_B=30$, the same calculation gives $2$. These examples assume a common target response and a common total-<protein> response between lanes, negligible remaining background, and comparable target recovery; they do not equate total-<protein> staining with target abundance. Multiplying both signals in a lane by the same loading factor cancels in their <ratio>; a selective target loss does not cancel.
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