How better antibodies can save time in the lab

Antibodies are among the most important protein-detecting reagents in research, yet they can be surprisingly unreliable. It’s an issue close to Alejandra Solache’s heart. “The way many antibodies are made and validated often isn’t as robust as it should be, and has created a lot of problems for researchers,” says Solache, senior vice president of research and development at Abcam, a global producer of research reagents.

Lack of specificity, and variability between batches, can produce misleading or inconsistent results and hamper progress. Such problems can undermine a study’s conclusions, leading to retractions or even product withdrawals².

Most antibodies used by researchers are either polyclonal — a mixture of antibodies derived from the serum of immunized animals — or monoclonal, where a single specific clone comprises the entire antibody pool. Polyclonals are prone to batch-to-batch variability when different animals are re-immunized, while the expression of monoclonals in hybridomas can vary over time, reducing specificity³.

In 2015, Andrew Bradbury, chief scientific officer at antibody engineering company Specifica, wrote a commentary in Nature calling for researchers to use recombinant antibodies to improve reproducibility⁴. The article was a wake-up call for research reagent firms. “There has definitely been a move away from polyclonals and traditional monoclonal antibodies towards recombinant ones,” Bradbury notes. But is their value truly appreciated?

The benefits of recombinants

Recombinant antibodies prevent many of the problems associated with polyclonal and traditional monoclonal antibodies. “If researchers knew the extent of the inherent variability of these antibodies, they would quickly switch to antibodies produced recombinantly,” Solache says.

Because recombinant antibodies are made in vitro by cloning specific antibody genes into vectors, their expression is controlled, improving consistency and reproducibility. Moreover, as the gene sequence is known, it can be used time and again. “Knowing that your antibody is going to behave the same way years down the line, gives you peace of mind,” Solache adds.

An added benefit is that recombinant antibodies can be tailored to researchers’ needs. They can be genetically engineered to improve affinity, functionality or specificity, or combined to form a ‘cocktail’ against different parts of the same target. These approaches are useful when working with proteins that are difficult to detect because of their structure, sequence or low expression levels.

There is, of course, another advantage. “Recombinant in vitro production using antibody libraries and modern DNA technology, instead of laboratory animals and hybridoma cell lines, is particularly attractive for customers who are steering away from animal-derived products,” Bradbury adds.

Validation at every step

Even with these superior reagents, robust validation and quality control is essential. “At Abcam, we put in an enormous amount of work into confirming the specificity of the antibodies,” Solache explains.

The company uses knock-out cell lines to eliminate doubts about cross reactivity, and has recently introduced biophysical quality-control methods, such as mass spectroscopy, high-performance liquid chromatography, and dynamic light scattering, to determine identity and purity, and identify potential aggregation, which could compromise activity.

Abcam offers more than 4,500 knock-out-validated recombinant antibodies, and is working with organizations all over the world to develop supplies of consistent, high-quality reagents. These antibodies are being used to improve understanding of many diseases, including young- onset Parkinson’s disease⁵, acute lung disease⁶ and cancer⁷, and to uncover potential therapeutic targets.

As recombinant technologies become faster and cheaper, Bradbury reckons they will take over as the primary production method. “If antibody therapeutics are produced this way, why not use it for your reagents? It’ll be faster and more reproducible. And if scientists can use the exact same reagents, it’s likely to reduce the number of retractions.”

References

<sub>1. Cox, T.R. et al. Nature 617, 208 (2023).
2. Fonseca, S.G. et al. Nature Cell Biol 17, 105 (2015).
3. Bradbury, A. R. M. et al. mAbs 10, 539–546 (2018).
4. Bradbury, A. & Plückthun, A. Nature 518, 27–29 (2015).
5. Laperle, A.H. et al. Nature Med 26, 289-299 (2020).
6. Hsu, C.G. et al. Cell Death Differ 29, 1790-1803 (2022).
7. Yuan, L. et al. Cell Death Differ 29, 1513-1527 (2022).</sub>