Choosing antibodies against tumour surface targets by epitope and application: why a her2 antibody and an erbb2 antibody are the same target under two names and why an erbb4 antibody is not a substitute, what a met antibody or c-met antibody, a mer antibody and a mertk antibody demand of receptor tyrosine kinase detection, how a ror1 antibody, a dll3 antibody, a steap1 antibody, a gprc5d antibody, a mesothelin antibody, an il13ra2 antibody, a caix antibody, a cd33 antibody, a cd37 antibody, a muc1 antibody, an l1cam antibody, a claudin 18.2 antibody, a pd1 antibody and a tissue factor antibody differ in whether the epitope survives fixation, and why a reagent used to score expression has to be validated against the assay it will inform rather than against a lysate

Choosing for the question

1. Establish which name the target goes by. Several of these targets have two or three names in common use, and catalogues index them differently. Confirm the gene identity before comparing reagents, because apparently different products are frequently the same target.
2. Match the epitope to the application. An extracellular epitope is what a live cell stain or a flow panel needs. An intracellular epitope may be the only one that survives fixation for tissue work. A reagent validated on lysate says nothing about either.
3. Treat scoring as a quantitative assay. If a stain will be used to grade expression, it needs controls spanning the range, a defined scoring rule and evidence that intensity tracks amount. Reagents sold for research are not calibrated for this.
4. Watch for shed and soluble forms. Several surface targets are shed, so a soluble measurement and a cell surface measurement can diverge. State which you measured and do not treat them as the same quantity.
5. Control expression, not just staining. A cell line panel spanning negative, low and high expression is the control that makes a stain interpretable, and it transfers across experiments better than any isotype control.

Presence and amount are different claims

Detecting a target is straightforward with a validated reagent. Grading how much is present requires a calibrated assay, a defined scoring rule and controls across the range, and the difference matters because expression level frequently decides whether a target is actionable.

Where a project will eventually need grading, build the control panel early. Retrofitting quantitation onto a stain designed to show presence is rarely convincing.

Two reagents, one conclusion

For any target whose expression will drive a decision, two independent antibodies against different epitopes agreeing is far stronger evidence than one reagent with a good datasheet. Disagreement is equally useful and better discovered early.

It costs one additional reagent and one experiment, and it is the standard a careful reviewer applies.

Common questions

Can a research antibody be used to score expression?

For internal research comparisons with proper controls, yes. For anything informing a clinical decision it cannot, because scoring reagents are validated and labelled for that purpose and research reagents explicitly are not.

Why does a reagent work on a blot and not on tissue?

Different epitope accessibility. Denatured protein on a membrane presents regions that a cross linked tissue section hides, and retrieval has to be optimised per reagent rather than per tissue.

Extracellular or intracellular epitope?

Extracellular for live cells, flow cytometry and anything measuring surface density. Intracellular where fixation has destroyed the surface epitope, accepting that the measurement is then total rather than surface protein.

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