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Finding the right antibody pair is a bottleneck in ELISA and lateral flow assay development. This article will walk you through the ins and outs of developing antibody pairs and help you formulate the most fitting plan for your assay development needs. While this article focuses on addressing challenges of IVD antibody pairs development, it also offers one useful tip for research grade (RUO) ELISA development. Enjoy reading (about 7 minute read).
Antibodies and standard proteins are the critical reagents in ELISA, lateral flow and many other IVD assays. Finding the antibody pairs to work in your assay is difficult.
Ideally, assay developers prefer to source commercial antibodies that pair well together. The challenge is, antibodies that work well individually often do not work well together. This is especially true for less popular biomarkers for which the antibody selection on the market is less abundant.
If the assay developer use custom developed antibody pairs, planning ahead and choosing the best fitting antibody discovery strategy is crucial.
Antigens tend to have some epitopes with higher immunogenicity than the rest. Immune reactions against these antigens tend to focus on these epitopes and not against the weaker epitopes. This way the majority of the clones on the market are against epitopes that are close together, and blocking each other from binding as a pair.
This is only one of many reasons antibodies do not work in pairs. The best chance of getting pairable antibodies is to screen them specifically for pairing purposes during development.
Identifying or developing these critical reagents can easily cost hundreds of thousands of dollars.
Both sourcing commercially available antibodies and creating custom antibodies for pairing are risky. It takes a lot of time and money to buy and validate antibodies for pairing, and there is no guarantee they will work for your application.
Conventional monoclonal technologies are not tailor fitted for making antibody pairs and are subject to the immune reaction bias described above (why is paring antibodies difficult). This makes the custom antibody option also risky.
Because of this, assay developing companies usually require their client to provide critical reagents.
While it is not the main topic of this article (IVD antibody pairs development), we will give this tip for developing RUO ELISA kits: Boster Bio has an ELISA platform that can turn a single polyclonal antibody and a protein standard into a custom made functional ELISA kit on our 90-minute Quick ELISA kit platform. For less than $5000, you can have 10 ELISA kits made for your target of interest without needing a pair of antibodies.
Finding one well validated polyclonal antibody is magnitudes easier than finding a pair. If you are looking to develop some RUO ELISA kits, contact us and find out how we can help today.
Note: this method does not fit IVD assays as you will never get FDA approval with a polyclonal antibody based kit. For research though it is sufficient.
Boster's Plasma Cell Discovery (PCD) platform is uniquely capable of generating antibody pairs. It is a recombinant rabbit monoclonal antibody discovery platform featured with the ability to screen entire splenocyte populations. You can read more about our custom rabbit monoclonal antibody service here.
As described before, the main challenge of finding good antibody pairs is finding a good second antibody that reacts with the antigen with high affinity and reacting to an epitope without interfering the first antibody binding to the antigen.
Boster's PCD platform excels at screening cells at large scale and discovering the rare population of cells with the highest affinity and specificity. It can also screen B cell clones using the reaction condition of the end application.
The antibody pair discovery solution starts with the regular rabbit monoclonal antibody discovery. Upon successfully finding the first antibody clone that reacts with the antigen, we can use the antigen proteins blocked by the first antibody to screen for the second antibody. This way any positive clone in the second round of screening is ensured to react against the antigen bound to the first antibody.
So a few other platforms can screen clones with antibody bound antigens. How is our PCD platform better suited for doing this? It all comes down to its ability to screen the entire spleen and finding the rare most splenocyte population with the highest affinity. In conventional situations this might be an overkill, as you only need a clone that is good enough. However in situations with special requirements, such as antibody pairing, PTM specificity, recognizing a point mutation etc. The condition is very specific, and the percentage of fitting clones decreases drastically. It is in these "tricky" situations that our ability to screen whole spleens make the biggest difference.
Read more about it here and book a free consultation and ask about our special antibody pair discovery package. This one-of-a-kind package is the only one on the market custom tailored to making antibody pairs for IVD assays, such as ELISA and lateral flow.