More than two years after the COVID-19 pandemic, scientists are still wracking their brains over a fundamental question: Is there anything they could measure to determine if people are protected?

Researchers call such measures “correlates of protection” – indicators that a person is unlikely to become seriously ill if they have contracted a pathogen such as SARS-CoV-2, the virus that causes COVID-19.

Previous data have focused heavily on a single parameter: neutralizing antibodies. These specialized proteins, produced by white blood cells called B cells after people become infected or vaccinated, help ward off future diseases by blocking viruses from entering cells.

This protection is short-lived. Antibody levels begin to decline after a few months. But “we’re not seeing hospitalizations increasing as fast as antibodies are declining,” says immunologist E. John Wherry of the University of Pennsylvania. “So what is it?”

A key source of protection for those previously infected, he and other experts suggest, are memory T cells. These persistent immune cells don’t necessarily fight off infection, but they do prevent mild symptoms from getting worse. They do this by recognizing parts of a virus or other invaders and activating processes that help other immune cells or destroy infected cells.

Given their persistence, T-cell levels measured in large numbers of people after infection or vaccination could help determine a protective correlate. Indeed, on April 21, Wherry and dozens of other researchers, physicians, and biotech officials sent a letter to the US Food and Drug Administration urging the agency to do so Monitor T cell abundance— alongside antibody levels — to better assess immunity at the city, state, or national level to determine the effectiveness of new vaccines being reviewed by regulators.

“The need for many doses is impractical at the population level. We need vaccines that provide broader protection and last longer, and there is still work to be done,” Ofer Levy, one of the signatories to the letter, said at a recent Harvard Medical School press conference. Levy directs the Precision Vaccines Program at Boston Children’s Hospital and is a member of the FDA’s Vaccines & Related Biologics Products Advisory Committee (VRBPAC).

Unlike neutralizing antibodies, T cells recognize a wide range of targets on the virus. While antibodies attach to small fragments of a single protein – the protruding “spikes” on the surface of SARS-CoV-2 – T cells not only recognize spike snippets, but a variety of other viral proteins. Each segment is presented on the cell surface by a set of scaffolding molecules called human leukocyte antigens (HLAs), which differ from individual to individual. (HLA typing is used to match patients and donors for blood or bone marrow transplants.)

As a result, each person’s T cells “see” spike protein segments differently because they’re held up by different scaffold proteins, says Brianne Barker, an immunologist at Drew University. An advantage of this scaffolding process is that it makes T cells less susceptible to viral evolution. Although some of the viral protein fragments targeted by T cells evolved to evade immune attack, others remain unchanged as targets. When a virus is passed to another person, their T cells can look for these unmutated snippets. “If the virus is trying to evade your T-cells and then it’s transmitted to me, all those evasive maneuvers and all the evolution it’s done doesn’t make sense because my immune system is signaling a different target,” Barker says. “It’s really difficult for a virus to evolve around T cells.”

Research confirms this. In a study described in a January Cell article, scientists at the La Jolla Institute for Immunology analyzed blood from 96 adults at various times after receiving a COVID vaccine. Six months after vaccination, neutralizing antibody levels had fallen significantly, while T-cell responses remained strong even against Omicron.

Additionally, in another study, people showed evidence of high-quality T-cell memory regardless of how often they were exposed to the virus through vaccination or infection. And the T cells didn’t succumb to “exhaustion” – a dysfunctional condition that can occur with chronic stimulation and that some scientists feared might occur with repeated vaccinations. These results were published on April 5th nature immunology.

Still, T-cell research needs more emphasis because, despite what has been shown so far, few studies have directly proven that T-cells help protect against COVID. “We all think they are, but it’s actually very hard to show,” says Paul Thomas, an immunologist at St. Jude Children’s Research Hospital in Memphis, Tennessee, who led the study nature immunology Research.

In one study, scientists collected blood plasma from a number of macaques they had infected with SARS-CoV-2 and found that infusing the plasma into naïve animals helped them resist later infection. When the researchers removed T cells from the plasma before the transfer, those recipients fared worse. Evidence for protective T cells also comes from small human studies, where cancer patients with impaired antibody responses had better survival rates when they had larger numbers of T cells.

But immunologists need more answers. If memory T cells help protect the immune system, how long do they stay in the bloodstream? How many are needed to stave off a serious illness? “We don’t know the answers to any of these questions because we don’t measure memory T cells sufficiently in enough patients,” says Wherry.

Even without large-scale surveillance, researchers have been examining patients’ T cells throughout the pandemic. Wherry’s lab, for example, has been doing in-depth analyzes of immune responses in 60 to 80 vaccinated people for more than a year – but only a handful have gotten breakthrough infections. The data are intriguing, but there’s “no statistical power as to whether they got infected because their T-cells were low or because they engaged in risky behaviors, like going to bars every night,” he says.

Getting those answers would require tracking thousands to tens of thousands of people, Wherry says. With this larger cohort, analyzes in each individual could be much easier – researchers could figure out how many SARS-CoV-2 specific T cells people have and where they fall in a range of T cell measures.

A major challenge in further development is of a technical nature: T cells are much more difficult to study than antibodies. Standardized assays can measure SARS-CoV-2-specific antibodies in tens of thousands of blood samples per day, and the process can be automated with robots. T cells must first be purified from blood samples – a procedure that requires several hours of work by a laboratory technician – then the cells must be cultured, stimulated with SARS-CoV-2 peptides and secreted molecules measured. T-cell protocols “are a lot more complicated,” says John Altman, an immunologist at Emory University. Doing the analysis on a large scale “would be enormously expensive, labor intensive and difficult to control and standardize across different sites”.

The main motivation for measuring T-cell responses “is to guide decision-making to improve vaccines,” says Altman. “We don’t need the data to give us better ideas about what to do. We already have reasonable ideas of what to do and we should just do them.” In one nature immunology In his comment last month, Altman suggested boosting immune memory by expanding the number of potential SARS-CoV-2 targets the immune system is ready to fight. This could be achieved by including additional nonspike antigens in future vaccine formulations to stimulate an immune response – a strategy already being implemented with T-cell priming peptide vaccines.

In fact, researchers are working on a streamlined T cell testing procedure. Measuring T cells is difficult, Wherry says, but he outlined several new approaches in a March 24 article science immunology point of view article. One is an assay that bypasses tedious cell purification and manipulation by detecting activated T cells in blood samples injected into tubes premixed with chunks of SARS-CoV-2 proteins. Another approach, which is more expensive but could be more easily scaled up, uses DNA sequencing to detect SARS-CoV-2 specific T cells in whole blood samples, rather than the more cumbersome methods of measuring the molecules derived from purified T cells. Cells are secreted into tubes or plates.

The letter to the FDA received a “huge response” from many, who agreed that T-cell measurements should be essential for future vaccine studies, Wherry says. “It was one of the most homogenous types of reactions I’ve seen given how polarized we are these days. I think there is a certain dynamic.”