Considering the recent waves of infectious diseases and the emergence of mRNA vaccines, it has never been more important to have a robust and rigorous means of measuring immune response. Newly circulating pathogens must be identified and characterized, which includes assessing the human immune reaction to infection. Furthermore, as mRNA-based technologies offer a pathway for much faster vaccine development, it is essential to understand how each patient responds to them as quickly as possible.
This need to measure immune response to infectious disease and related vaccines may be more pressing than ever, but it’s certainly not new. Since the 1970s, the workhorse technique used in most laboratories has been the enzyme-linked immunosorbent assay (ELISA) – popular thanks to its efficiency and versatility. It can be conducted at high capacity, usually in 96-well plates, with results available within a day. ELISAs are also able to bind various analytes, which enables testing for different antigens as required.
However, widespread use of a tool does not qualify it as the ideal. ELISAs can only test for one immunoglobulin isotype or subtype for a single antigen at a time – yet modern immune analysis requires the interrogation of several parameters at once. Just imagine how limited the COVID-19 public health response would have been had scientists not been able to easily test for antibodies to three different proteins to distinguish between the immune response from a vaccine versus a natural infection. With ELISAs, a test must be run for each antibody isotype or subtype of each antigen, with a new biological sample every time – and that’s simply not feasible for the comprehensive immune response analysis required today.
More scientists have turned to a bead-based multiplexing technology that allows for comprehensive measurement of the immune response by testing for as many as 500 analytes – including antibodies, cytokines, chemokines, or nucleic acids – from a single sample. This approach allows for rapid results, generating information in just a few hours.
Bead-based multiplexing has been demonstrated to help quickly characterize emerging infectious diseases and assess the immune response triggered by mRNA and other types of vaccines.
Emerging diseases
Having a rapid, multiplexed approach for in-depth analysis can make a difference in the scientific and public health response to emerging infectious diseases. For example, researchers in the Democratic Republic of the Congo and France have used this approach to better understand the spread of mpox virus in humans. This disease has hit the Democratic Republic of the Congo particularly hard, and researchers were concerned that incidence metrics could be skewed. Why? Because clinical case reporting tends to overestimate the rates of infection, while lab-confirmed PCR results can underestimate them.
For a more realistic assessment of the toll that mpox was taking in the country, the researchers performed antibody testing, which can better assess infection rates and identify transmission chains during an outbreak (1). By implementing bead-based multiplexing technology, they were able to generate substantial information from residual blood samples left over after clinical testing had been performed.
The study included samples from 463 patients collected during nearly 175 separate mpox outbreaks from 2013 to 2022. Data from PCR testing was available for comparative purposes. This approach not only gave the team deeper insight into the samples that had tested positive before (they were able to look for antibodies to at least three mpox-associated peptide antigens), but also allowed them to correct previous misclassifications. Samples from a total of 66 patients were found to be positive after being missed by PCR-only results, presumably because those PCR tests were run at the wrong time to detect infection. This increased the known mpox infection rate in the cohort from 34 percent to 48 percent, giving the team a clearer view of the epidemiological landscape and allowing them to spot infections in regions previously not known to harbor outbreaks.
Bead-based multiplexing has been successful for several other infectious diseases. In one recent report, scientists deployed bead-based multiplexing technology to evaluate the utility of tracking antibodies for 20 peptides associated with serotypes of dengue fever, Zika virus, yellow fever virus, chikungunya virus, and West Nile virus, as well as previous vaccinations to other viruses (2). The ability to examine so many analytes in hundreds of samples made it possible to identify the peptides that would be most useful for serology screening in areas with high incidences of flavivirus infections. A similar method was used to distinguish between active and latent tuberculosis infections using more than 200 antigens (3).
Vaccine response
Teams around the world used bead-based multiplexing for diagnostic and serology assays to track COVID-19 infection rates and paths (4). However, the pandemic also provided an opportunity to extend the use of this technology to vaccine evaluations – an approach that has previously been used successfully with vaccines for human papillomavirus, Streptococcus pneumoniae, and many other infectious diseases.
Though bead-based multiplexing was used in the development of the original Pfizer/BioNTech mRNA vaccine for SARS-CoV-2, it has been more commonly deployed for assessing the immune response to vaccinations and comparing that to the protection acquired from a natural infection (5). For those analyses, multiplexing capabilities are essential (we must test for the antigens included in vaccines, as well as the additional antigens specific to the virus itself).
One group of researchers used bead-based multiplexing technology to measure the humoral immune response to two different mRNA vaccines for SARS-CoV-2 in both humans and rhesus macaque models, with the analysis extending for months to include two separate doses of each vaccine (6). They tested plasma IgG levels against seven different coronaviruses and examined the neutralization capacity of antibodies found in plasma. The study identified specific IgG subtypes associated with vaccine response and found higher IgG levels elicited by the Moderna vaccine compared with the Pfizer vaccine.
In a separate study, scientists assessed IgG responses in standard serum samples and nasal fluids to understand the immune protection achieved through nasal mucosal antibodies (7). They determined that, though vaccines trigger strong immune responses in general, natural infections lead to higher levels of mucosal antibodies in the nasal passages, where defenses might be most important. The team suggested that future vaccine development should focus on approaches that might yield greater levels of mucosal antibodies.
One final example comes from a study inspired by concerns that the use of monoclonal antibody treatment for SARS-CoV-2 patients could weaken future immune response to vaccines (8). Researchers conducted a six-month serology survey using bead-based multiplexing technology to evaluate vaccine response among 135 nursing home residents and staffers, including patients who had previously received monoclonal antibody treatment. Results indicated that exposure to the monoclonal antibody had no effect on the immune response triggered by vaccination, alleviating fears about the dangers of endogenous immune response to therapy.
These few studies reflect just a small snapshot of the scientific and clinical explorations conducted during COVID-19, but they show how thoroughly researchers can interrogate immune response to vaccination using bead-based multiplexing tools.
And though COVID-19 is now less of a focal point, there are many other cases of emerging infectious diseases – with associated vaccine development efforts. For a comprehensive view of the immune response to natural infections and vaccines, bead-based multiplexing offers significant advantages – in time, cost, and analytical depth – compared with traditional methods, such as ELISAs.
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References
- E Kinganda-Lusamaki et al., Pathogens, 12, 7 (2023). PMID: 37513764.
- F Falconi-Agapito et al., Front Immunol, 13 (2022). PMID: 35154111.
- N Nadege et al., Front Immunol, 13 (2022). PMID: 35514994.
- A Cameron et al., Microbiol Spectr (2022). PMID: 35389244.
- MJ Mulligan et al., Nature, 590 (2021). PMID: 33469216.
- R Ravindran et al., PLoS One (2023). PMID: 37856429.
- O Puhach et al., EBioMedicine, 98 (2023). PMID: 38035462.
- RJ Benschop et al., Sci Transl Med, 14, 655 (2022). PMID: 35679357.
