HOW TO CHOOSE THE RIGHT ELISA KIT

The enzyme-linked immunosorbent assay (ELISA or EIA) is a laboratory method to detect and quantify the presence of a protein in biological samples (1, 2).

When selecting an ELISA kit, researchers are often confronted with the question which assay to choose of the many commercially available kits.

It can be a challenge! Here are a few hints that may help.

HOW TO CHOOSE THE RIGHT ELISA KIT

As a general rule, before purchasing an assay, always read the protocol booklet (instructions for use – IFU or package insert) in detail. This should ensure that the kit will be suitable for your requirements. Check out the following:

1. ANALYTE

Which protein biomarker will you be measuring? Be sure to use the correct term during your search. Some biomarkers have alternative names (e.g. Sclerostin or SOST ELISA (SOST is actually the name for the gene that encodes Sclerostin).

2. SPECIES – SPECIFICITY – CROSS REACTIVITY

Verify if the assay can be used in the respective model such as e.g. human, rat, or mouse. Due to high homology between species, some ELISA kits work both in humans and in different animal species. As an example the biomarker ELISA kit for NT-proANP was developed for human use but due to the high sequence homology between species, the kit is successfully used to measure NT-proANP as a cardiac safety biomarker in various animal models (rat, mouse, rabbit, monkey).

3. SAMPLE TYPE

What is the sample type (matrix) you´ll be using (e.g. serum, EDTA-plasma, heparin-plasma, citrate-plasma, cell culture supernatants, urine..) ?

Verify if the assay is compatible for your sample type: check the package insert and, if available, check if there are validation data showing results (often found on the manufacturers website).

Of note: analysis of some biomarkers in the “wrong” matrix can lead to “false” results due to a matrix effect.

4. SAMPLE VOLUME

Check the amount of sample required per well (calculate volume to measure your samples in duplicates). Low sample volumes and precious samples are often a selection criterium.  

5. SENSITIVTY – BIOMARKER CONCENTRATIONS TO BE EXPECTED

Before choosing an assay, look into the validation data of the kit (often documented in the ELISA protocol booklet or in the validation data files).  

Reference values and pathological values in serum and/or plasma of the biomarker of interest are sometimes documented as well. These data can be helpful in selecting an appropriate assay. In some cases samples may require a pre-dilution. Therefore, always verify if the dilution buffer (assay buffer) is included in the kit or ask the assay developer for their input.

Of note: assays offering high sensitivity offer a different dynamic range than assays with a lower sensitivity. The dynamic range of an assay indicates the range of concentrations over which an assay can accurately quantify the analyte.

6. ASSAY PERFORMANCE – ASSAY VALIDATION

Careful evaluation of the assay´s performance characteristics is important in selecting an ELISA kit.

Choose an assay that has gone through a rigorous validation process. Check out if you can find data on the following performance characteristics:

• Accuracy- detection of a protein biomarker in clinical samples.
• Dilution linearity and parallelism  – recovery of the analyte of interest in diluted samples
• Specificity & cross-reactivity – making sure that you detect only the analyte of interest
• Precision – within-run and in-between run precision – ensuring precise and reproducible results within an across assay lots
• Calibration – ensures consistent performance over the range of the assay of the calibration curve
• Sample stability – ensures the stability of the analyte of interest (e.g. exposure of real samples to multiple freeze-thaw cycles, stability at room temperature..).

 

7. KIT COMPONENTS

Verify if the contents of the ELISA kit includes all the necessary components e.g. controls, assay dilution buffer. Consider storage requirements such as temperature sensitivity and expiration date.

8. CITATIONS & REFERENCES

Check if there are citations on the manufacturers website for the specific ELISA kit. Look into publications and seek feedback from researchers who have used the assay you are considering.

9. PRODUCT ORIGIN

Verify if the kit supplier is the kit manufacturer. More and more kits are repacked and are sold under different names, although it is always the same kit.

ELISA kit manufacturers will more likely give you qualified support as they “know” their product (e.g. availability of additional calibrators, controls, buffers.., technical know-how on the kit..).

10. CUSTOMER SUPPORT

Verify if the kit provider can provide timely and helpful customer service.

 

Related Literature

1.Enzyme-Linked Immunosorbent Assay: Types and Applications. Hayrapetyan H, Tran T, Tellez-Corrales E, Madiraju C. Methods Mol Biol. 2023;2612:1-17. doi: 10.1007/978-1-0716-2903-1_1. PMID: 36795355.

2. Enzyme Immunoassay (EIA)/Enzyme-Linked Immunosorbent Assay (ELISA). Lequin RM, Clinical Chemistry. 2005; 51, 12:  2415-2418.