Buying a tet3 antibody, a cyp26b1 antibody or a rab27b antibody: how to judge a reagent for a target almost nobody has benchmarked, and the validation evidence to demand before it decides an experiment

Judging the reagent before you buy

1. Read the immunogen, not the marketing. Which region of the protein, and is it shared with the family? Enzymes and regulators frequently sit in families with high similarity, so an antibody raised against a conserved domain will recognise the relatives too. If the immunogen sequence is not disclosed, that itself is information.
2. Check the applications claimed against the evidence shown. A datasheet listing six applications with one western blot image has evidence for one. Treat each application as separate: performance on a denatured blot says very little about performance on a fixed section or in a pulldown.
3. Insist on a genetic negative where one is possible. A knockout or knockdown sample is the only control that distinguishes specific binding from a band of the right size. Where no knockout exists, an overexpression sample is the next best thing and considerably better than nothing.
4. Prefer a recombinant monoclonal for a long project. For a target you will work on for years, a defined sequence removes lot variation entirely. A polyclonal against a niche target may be a single bleed, and when it is gone the project restarts its validation from the beginning.
5. Buy one lot, and bank it. Ask the supplier how much of the lot remains before committing. For a low-demand target the answer is often small, and discovering that halfway through a study is the expensive version of this conversation.

Recording what you did, so the next person does not repeat it

Catalogue number, lot, dilution, application, sample, and the control that was run, kept with the result. For a niche target your own record becomes the literature, and a year later you will be the person asking whether this reagent was ever properly tested.

Publish the validation images as supplementary material where you can. Antibody validation that stays in a notebook is validation the field pays for again.

When no usable antibody exists

Epitope tagging the endogenous locus removes the problem entirely: a well characterised tag antibody works the same way on every target. It costs an editing project and it is frequently faster than screening six antibodies that do not work.

Where the readout can be transcript rather than protein, quantitative PCR or sequencing avoids the reagent problem altogether. It answers a different question, and it is worth being explicit about which question the experiment actually needs answered.

Buying from a smaller supplier

Niche targets are often served by smaller specialists, and the useful questions are whether the antibody is made in house or resold, what the lot history is, and whether they will supply a small trial size. A supplier who resells cannot answer questions about production.

Ask for a trial size before the full vial. Suppliers who believe in a reagent generally agree; the ones who refuse have told you something useful.

Common questions

How do I validate an antibody nobody has published on?

With a genetic negative if one exists, an overexpression positive if it does not, a size check against the predicted mass with the modifications you expect, and a second antibody against a different epitope. Agreement between two independent antibodies is the strongest evidence available without a knockout.

Is a polyclonal or a recombinant monoclonal better for a niche target?

A polyclonal is often more sensitive and is finite. A recombinant monoclonal is reproducible indefinitely and can be reformatted. For a multi-year project the recombinant is usually the cheaper decision by the end.

What if the antibody detects a band of the wrong size?

Check the predicted mass against known isoforms and modifications before concluding anything, then run the genetic control. A band at an unexpected size that disappears in a knockout is real biology; one that does not is the antibody.

Does a supplier's validation badge mean the antibody is validated?

It means the supplier applied its own scheme, and those schemes differ a great deal. Read what was actually done and in which application, and treat it as a starting point for your own control rather than as a substitute.

Browse by service class

• Compare Gene synthesis on advertised prices