Chemiluminescence is a process in which a chemical reaction produces light as a byproduct. In immunochemistry, it is used in assay and detection systems for techniques such as CLIA (Chemiluminescent Immunoassays).
Researchers use CLIA for gene expression studies, protein quantification, and immunoassays, among other applications. For this purpose, most researchers use a CLIA kit.
This kit includes a complete set of reagents and components for performing a chemiluminescent immunoassay.
However, you cannot simply purchase a CLIA kit from any vendor for your experiments. It needs to be QC tested, certified by ISO, and have accurate sensitivity.
There are many other markers to check before making a purchase. Let us discuss that ahead in the blog. Stay tuned.
1. Species Reactivity
The whole point of buying a CLIA kit is to fulfil the purpose of your experiment, so the first step is to confirm whether the kit reacts with the species you are studying. Some Chemiluminescent Immunoassay (CLIA) kits are specific for human proteins, while others may work with mouse, rat, or other species.
If you ignore this step, chances are your antibodies won’t bind or generate no light signal. Wrong species reactivity will waste both the valuable biological sample and the reagent.
Note: Don’t fall for the ‘this might do’ trap. Be specific when you choose the Kit for your research.
2. Specificity
The second thing to check is the kit specificity. For example, if you are buying a Human COL2α1 CLIA Kit, the specificity must be defined for the natural and recombinant forms of COL2α1.
There must be no cross-reactivity between the Human COL2α1 and analogues. This ensures your kit won’t detect other molecules that are similar to COL2α1 and won’t give you a false positive.
3. Sensitivity
Next, check the sensitivity of the kit, which tells you the smallest concentration it can detect. A kit with low sensitivity cannot pick up small changes in protein levels. This is a problem if your study requires the detection of biomarkers present in tiny amounts.
Without the right sensitivity, the results may appear as if your target protein is absent when it is actually there in trace amounts.
Example: The sensitivity value for COL2α1 (Collagen Type II Alpha 1) (COL2a1): Min: 75 pg/mL; Max: 8000 pg/mL.
It indicates that this kit can detect as low as 75 picograms per milliliter of COL2α1 in your sample. A concentration of 75 pg/mL is a very low level, indicating the kit’s high sensitivity and its capability of detecting even small amounts of the target molecule in a sample.
4. Assay Type
Ensure that the type of CLIA assay is specified. Many Chemiluminescent Immunoassay (CLIA) Kits use a sandwich format, where the protein is captured by one antibody and detected by another.
Others use a competitive format where the target antibody is attached only to the antigen, followed by luminol and a catalyst. Using the incorrect assay type for your experiment can yield inaccurate concentration readings.
A general rule is that the sandwich assays are suitable for large proteins, while competitive assays are used for smaller molecules.
5. ISO Certification
You will also need to check if the kit is manufactured in an ISO-certified laboratory. ISO certification ensures quality standards in production.
Without certification, there is a risk of inconsistent reagent quality, contamination, or variation between batches. This can lead to experiments that are not reproducible and data that other researchers cannot trust.
6. Assay Application
Kits are designed for specific applications such as serum tests, plasma, tissue lysates, or cell culture supernatants. If you run samples outside the validated application, the assay might not work.
For example, hemolyzed blood may give false signals. Always check applications in the product description to avoid invalid results.
7. Quality Control Testing (QC Testing)
Confirm if the kit has passed QC checks for repeatability, recovery, and cross-reactivity. QC testing ensures that each batch is tested for precision.
Chemiluminescent Immunoassay (CLIA) Kits without QC testing may show high variation between wells or between assays. This can lead to inconsistent results and require you to repeat the experiment.
8. Delivery and Storage Instructions
Inspect how the kit will be shipped. Many CLIA reagents must arrive cold or on dry ice. If shipping conditions are broken, enzyme activity and antibody binding may be lost.
Also, ensure that you have the correct storage conditions in your lab for the kit. Some require temperatures as low as -20°C, while others may only need to be maintained at 4°C. Incorrect storage will reduce reagent stability and weaken the light signal, resulting in inaccurate assay results.
Conclusion:
Chemiluminescent Immunoassay (CLIA) Kits are important tools that give reliable and sensitive results when chosen and handled correctly. However, if you overlook basic checks such as species reactivity, assay type, or storage instructions, you may encounter failed experiments and wasted samples.
Always review the manual, confirm batch quality, and store the kit properly after delivery. Pay attention to the factors mentioned above, and your CLIA experiments will produce strong, reproducible, and trustworthy data.
