Mouse IL-6 ELISA kit and IgG kit selection: why species and range decide it

Choosing the right kit

1. Match the species exactly, every time. Cytokines are less conserved than their shared names suggest, and a kit raised against the mouse protein generally will not detect the human one. Check the species on the kit against the species of your sample before anything else, because this is the error that wastes a whole plate.
2. For immunoglobulin kits, check the subclass and the specificity. A total immunoglobulin kit and a subclass-specific one answer different questions, and an anti-species reagent may or may not distinguish subclasses. For antibody quantitation in a culture supernatant the subclass usually matters.
3. Match the range to the expected concentration. Immunoglobulin in a supernatant and a cytokine in serum sit orders of magnitude apart. Pick the kit whose standard curve brackets your expected values near its middle, where precision is best, rather than diluting to fit a convenient kit.
4. Qualify in your own matrix. Spike recovery and dilution linearity in the actual sample type. A kit validated in serum used on culture medium, lysate or a tissue extract is unvalidated until you demonstrate the matrix is not interfering.
5. Fix handling before the first plate. Tube type, time to centrifugation, storage temperature and freeze-thaw all move immunoassay results, and several cytokines are released during clotting. Write the protocol down and apply it to every sample in the study.

Running the plate so the numbers survive review, whichever IFN gamma ELISA kit you use

Randomise samples across the plate rather than loading groups in blocks, run standards and samples in duplicate and look at the duplicates rather than only the mean, and put a bridging control on every plate in a multi-plate study.

Fit the curve with the model the kit specifies and treat values read from the flat ends of it as far less certain than the coefficient of variation suggests.

Kit against a custom assay

A kit arrives qualified and costs more per sample. A custom assay from a matched antibody pair is cheaper at volume, takes real development time and gives control of supply, which matters over a multi-year study.

Whichever route, bank one lot of the critical reagents and the standard for the whole study. A new standard lot shifts the entire curve and that shift is indistinguishable from an effect.

Mouse IFN gamma, and the other mouse cytokine plates

Mouse IFN gamma is the readout most Th1 and vaccine studies end on, and its plate behaves like the other mouse cytokine kits with one caveat: the working range is narrow and stimulated supernatants routinely run off the top of it, so the dilution series matters more than the antibody pair. Buy the kit matched to the matrix you actually have, serum, plasma or culture supernatant, because the calibrator diluent differs between them. Where several cytokines are wanted from one small sample, price a multiplex plate against the sum of the singles before ordering four kits.

An il1b elisa kit, and the form it detects

Interleukin 1 beta is made as a precursor and cleaved by caspase-1 to the active cytokine, and a kit may recognise the pro-form, the mature form or both, which is why two kits can disagree on the same supernatant. Read the antibody pair's stated specificity before the sensitivity figure. Sample handling matters too: the cytokine is released on cell lysis, so a haemolysed or roughly handled sample reads high, and the working range for a stimulated supernatant usually needs a dilution series rather than a single point.

purified mouse igg, and what it is for

Purified immunoglobulin from non-immunised mice is a control reagent: an isotype-matched substitute for a primary antibody, which shows how much of a signal is Fc binding, non-specific adsorption or the detection system rather than the target. It also blocks Fc receptors before staining. The isotype and the subclass have to match the antibody being controlled, and the concentration has to match too, since an isotype control at a tenth of the antibody's concentration proves nothing about it.

An a20 antibody and a tnfaip3 antibody are one protein

A20 is the protein and TNFAIP3 is the gene, so an a20 antibody and a tnfaip3 antibody are the same target under two conventions and a search on one finds half the market. It is an inducible inhibitor of inflammatory signalling, so it reports a condition rather than a constant, and an unstimulated control is what makes a band readable.

A tnfaip3 antibody and the paralogue trap

Because the gene symbol resembles the unrelated TNFAIP genes and the protein's name resembles other A-numbered proteins, a tnfaip3 antibody has to be ordered against the gene identifier, and its own inducibility means the positive control is a stimulated lysate rather than a resting one. It is a ubiquitin editing enzyme, so substrate ubiquitination is the functional readout.

Common questions

Can a mouse ELISA kit measure the human protein?

Generally no. These proteins are less conserved than their shared names suggest, and the antibody pair is raised against one species. Check the species on the kit against your sample before ordering.

Total IgG or subclass-specific?

It depends on the question. For quantifying a monoclonal in a supernatant, the subclass-specific kit is usually the right one; for total immunoglobulin in a fluid, the total kit. They are not interchangeable and the standards differ.

Why does my kit saturate?

The range does not match the sample. Immunoglobulin concentrations sit far above cytokine ones, so a kit chosen by target name rather than by expected concentration will either saturate or read below its floor.

Do I have to qualify a commercial kit?

In your own matrix, yes. Spike recovery and dilution linearity take one plate and tell you whether the matrix suppresses or enhances the signal, which no datasheet can tell you about your samples.

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