Written in blood

In 1959, in a burgeoning riverside market town in what was then the Belgian Congo, a Bantu man fell ill.

An unknown virus was replicating furiously in the white blood cells circulating in his veins. A sample of his blood, taken for a study of the genetic diseases of red blood cells, was frozen and forgotten for nearly three decades. It contained traces of the mysterious virus that would take some 20 million lives worldwide by the year 2000: the human immunodeficiency virus (HIV), a retrovirus that thrives within the very cells meant to fight off invaders.

Blood is an ephemeral substance. Red blood cells, which make up nearly half of the blood’s volume, have a life span of just 120 days, give or take a week. Four months from now, every one of the 25 trillion or so bi-concave blood cells sailing through the body’s veins and arteries at this moment will have been consumed by macrophages and replaced by brand-new oxygen-toting cells. Outside the body, blood cells break down in 24 hours or less.

Frozen blood, on the other hand, is a liminal substance—neither alive nor dead. The Bantu man’s frozen blood became a snapshot of the past, and as medical technology and tests advanced, a resource for the future. Eventually, the Bantu man’s blood would help scientists trace the origins of one of humanity’s deadliest epidemics.

Not only can frozen blood be used to track the spread and evolution of the diseases that plague humans, but it can also transcend disciplines, providing valuable insights to geneticists, anthropologists, and immunologists alike. Within those blood cells are traces of humanity’s evolutionary history.

For more than four decades, blood has been the main focus of the work of Kenneth K. Kidd, Ph.D. A professor of genetics, of ecology and evolutionary biology, and of psychiatry at Yale, Kidd got his start in genetics at the University of Wisconsin, where he studied blood group typing to identify and register purebred cows.

In the 1980s, when population geneticists acquired the tools to look directly at DNA, Kidd and his wife, Judith R. Kidd, Ph.D. ’93, switched to human subjects. “We gradually moved more and more into population studies, just because they’re fascinating,” said Ken. The Kidds became interested in the genetic variations that occur between geographically distinct populations.

“There is just a terrific amount you can learn from the study of human populations,” said Judy, a retired research scientist in the Department of Genetics who earned her Ph.D. in anthropology. “When Ken was studying cattle genetics as a graduate student, I thought, ‘Oh, that’s interesting.’ But once he moved to humans, I thought, ‘Oh, that’s interesting!’ ”

Their work on genetic variation has helped to piece together the human genetic map. The Kidds have scoured the genome for genes involved in complex neuropsychiatric disorders, narrowing down genes that play a role in Tourette syndrome, schizophrenia, and alcoholism. They’ve also used DNA polymorphisms to trace human origins. Their research has shown that sub-Saharan African populations tend to have more variation in certain genes than other populations throughout the world, which could mean that the genes have had more time to accumulate mutations and diversify in those African groups. This work bolsters the theory that modern humans spilled out of Africa to populate the rest of the world.

The Kidds share all the data on genetic variation they collect in a publicly accessible bioinformatics database that they affectionately call ALFRED, short for the Allele Frequency Database. But the physical cell lines themselves are locked inside steel-colored freezers in their New Haven lab. As the Kidds wind down their careers, what will happen to their collection is an open question.

A “cold chain” to move blood

Cold storage is not new; insulated iceboxes have been around for at least 200 years. Your grandmother probably had a mechanical refrigerator to keep her perishables cool before one was ever used to chill blood. Mechanical refrigeration replaced consumer iceboxes in the 1930s—before that, commercial refrigerators were as likely to catch fire, explode, or leak toxic gas as they were to keep things cold—but the technology didn’t take off in the scientific world until World War II.

Soldiers scattered across the globe in need of blood transfusions led to the creation of a “cold chain” for blood transportation. This chain moved stateside blood donations to soldiers on the front lines, requiring not just fridges and freezers at collection sites, but also something to keep blood cold in transit: dry ice and liquid nitrogen.

Yale medical historian Joanna Radin, Ph.D., found that by the 1960s, scientists had reversed the flow of these cold channels, using the infrastructure devised during wartime to learn more about human biology. Blood collected from remote populations around the world could be shipped to high-tech laboratories for analysis, said Radin, an assistant professor in the history of medicine whose work examines the ways in which technologies of cold storage have transformed the practice of biomedicine and public health. This chain was not without kinks. In 1968, a World Health Organization (WHO) report advised researchers to scratch serial numbers directly onto the glass vials after several shipments of specimens were unidentifiable upon arrival at labs—presumably, the labels had washed off in melted ice.

The WHO, famous for its work on infectious diseases, was quick to recognize the potential benefits of cold storage. The agency saw the collection of serum, the liquid component of blood, as a way to expand the study of current diseases in remote parts of the globe and to prepare for the diseases of the future.

As new diseases—and tests for their detection—emerged, researchers could thaw frozen serum samples and study them in light of such medical advances.

The agency established four serum banks, repositories of frozen samples, to serve as a biomedical resource. The only one in the Western Hemisphere was opened at Yale in 1960. The WHO chose John Rodman Paul, M.D., to direct it.

Paul, chair of the Section of Epidemiology and Preventive Medicine at Yale, had used blood samples to study the spread of diseases like polio and was an early proponent of freezing biological specimens. In the 1940s, while investigating antibody patterns in Eskimos in northern Alaska, he froze serum samples so that he could reanalyze each one as tests for newly discovered antigens became available. After more than a decade of studying the same samples, Paul still found the frozen serum to be a fruitful source for epidemiological research. “Indeed,” he wrote in 1961, “the epidemiologic story which this work has gradually been unfolding is not yet finished.”

The reference serum bank at Yale, whose rise was the subject of Radin’s research, quickly became a hub for the advancement of epidemiological research. In 1966, Alfred S. Evans, M.D., M.P.H., professor of epidemiology, took over as the second director of the bank. In that year alone, more than 21,000 serum samples were handed out to scientists who wished to study them. Before the decade was out, the Yale collection had grown to over 25,000 individual samples. Yale could no longer house the substantial collection, and the bank moved to a building owned by the New Haven Cold Storage Co. By the 1990s, when the collection moved from Yale to the National Cancer Institute in Bethesda, it contained over 50,000 unique serum samples—and untold insights into the infectious and chronic diseases of the past and present.

A prescient WHO report released in 1959—the same year the Bantu man’s blood was collected and frozen—stated: “If samples of the sera collected in these surveys are stored in such a way as to preserve antibodies, it will be possible to examine them in the future and so to determine the past history of infections as yet unknown and to follow more clearly the changing pattern of communicable disease all over the world.”

Two decades later, HIV emerged as a devastating pathogen. In the early 1980s, young, otherwise healthy people the world over began to waste away, dying of infections that usually felled only the old: tuberculosis, bacterial pneumonia, cryptococcosis, and herpes among them. Eventually, researchers discovered that HIV was at the heart of this immune system failure. Scientists sought the origins of this viral invader, hoping that understanding where the virus came from might provide clues for a cure or, at the very least, a way to contain it. In 1998, researchers from The Rockefeller University tracked it down to a freezer at Emory University in Atlanta. The freezer held more than 1,000 blood samples collected in the Congo in 1959 for a study of an inherited blood disorder common among some Africans. Once HIV was isolated as the cause of AIDS, an Emory scientist told a colleague about those samples. Only one sample, belonging to a Bantu man from a riverside market village, tested positive for HIV antibodies, making it the oldest biological trace of the disease ever documented.

With this and other preserved HIV-positive tissues, researchers used the known mutation rate of the virus to approximate when the epidemic began. By 2008, scientists had estimated that HIV crossed over to humans from chimpanzees somewhere in southeastern Cameroon a century earlier, about 1908. And a greater understanding of both the origin of the virus and its mechanisms of action has contributed significantly to medical advances against the disease.

A question of ownership

The WHO didn’t advocate preserving biological samples just because they might come in handy in the future, Radin has shown; the agency was equally interested in preserving the past. Frozen blood samples could also serve as permanent genetic snapshots of indigenous populations that began disappearing almost as soon as they were discovered, and help to unravel humanity’s evolutionary history.

By the mid-20th century, globalization was heating up. The WHO envisioned a future of both environmental and cultural upheaval, in which lands and peoples would be overwhelmed by the spread of Western influences. According to Radin, the WHO considered so-called “primitive” island populations to be unique laboratories “in which nature had run experiments on humans for thousands of years.”

These populations provided unique windows into our evolutionary past, and according to the WHO, they were also vanishing. Scientists scrambled to collect blood samples from remote and endangered peoples to preserve their genetic fingerprints. Yale would become the home of a second massive collection, different from Paul’s bank of serum samples because it contained cell lines. Today, one of the largest collections of anthropologically relevant cell lines resides in the Kidds’s lab, according to Radin.

“Some of these samples are never going to be collectable again,” said Ken Kidd. Their collection contains samples from populations as diverse as the Atayal tribe in Taiwan to the Zaramo people of Tanzania. These isolated populations are not necessarily dying off, as WHO officials imagined they might, but rather integrating into societies that have grown up around them. Either way, says Ken Kidd, such change “obliterates a window into the past.”

Despite these pressures, many cultures expected to disappear are alive and well. But some have objected to the use of their genetic samples for research. “Cold War-era scientists’ vision of the future did not include a world in which the barriers to collecting such blood would come from these peoples themselves,” wrote Radin.

There are many reasons why a group would object to how their blood samples might be used. The Havasupai, a community that lives deep within carved cliffs of the Grand Canyon, is a prime example. In the 1960s, rates of diabetes among tribe members mysteriously skyrocketed. Thirty years later, a geneticist at Arizona State University in Phoenix parsed the tribe’s DNA for the cause. Roughly 100 tribe members volunteered blood samples, but when they later learned that those samples had been used for other studies, including one that proposed a theory of the tribe’s origins that contradicted the group’s own creation myth, they balked. After a lengthy legal battle, the university paid reparations and returned the blood samples.

Ten years after their cells had been immortalized in a cell line, another group of Native Americans, whom the Kidds prefer not to name for legal reasons, asked that their cells be returned to them even though they had originally given full informed consent. But because the process of transforming a cell into a cell line involves infecting it with a virus, those cells are designated a biohazard—the Kidds couldn’t ship them to anyone without a functioning lab and a qualified technician. The group settled for having the cell lines and DNA destroyed. “The head of the Human Investigation Committee here at Yale watched us thaw out and destroy all the frozen cell lines,” said Ken. The DNA that had been isolated from those cells was “cooked” in an autoclave.

“That was painful,” Ken said of the ordeal. “Oh, it was heartbreaking,” added Judy. Today, the Kidds’s entire collection faces an uncertain future.

Judy retired in 2011, and Ken is likely to retire by 2016. It’s unclear what will become of the cell lines they’ve spent their careers studying. “It’s a matter of interest and responsibility and money,” said Judy. “There are lots of things we could do that wouldn’t make me feel as good as keeping those cell lines right here at Yale.” Ideally, the collection will remain at Yale, perhaps in the Peabody Museum as a frozen gallery of sorts, where a technician can watch over the cells, monitor liquid nitrogen levels, and send cell lines to qualified researchers. The scientist or organization, at Yale or elsewhere, who takes responsibility for the cells will be responsible for ensuring both the immortality of the cells and the immortality of the promises that were made to donors when they gave their blood to research. If both needs aren’t met, all 2,600 lines will have to be destroyed.

If the cells remain in the freezers, they could yield untold knowledge for future generations of researchers. The issue remains, however—the needs of science may conflict with the social and ethical questions raised by the maintenance of cold blood. “They are not always likely to be in alignment,” Radin said. “Choices made in the past have consequences for the present, and what might have seemed ethical in one place and time will change.”

Kate Wheeling was Yale Medicine’s 2014 writing intern.