Campbell, Pressman, Pauling and the Binding of Antibodies

Drawings of the interaction between an antibody and azoprotein by Linus Pauling. 1940s.

Dan Campbell first collaborated with Linus Pauling on a fellowship at Caltech in 1940, during which time the duo tried to explain how antibodies are formed. At the time, Pauling believed that antibodies were proteins in-the-making that needed to bind to antigens in order to fold and complete their structure. If this principle was correct, Pauling thought, it might be possible to create artificial antibodies by simply denaturing proteins and allowing them to bind and refold in the presence of antigens.

Despite the fact that Campbell’s initial test results cast doubt on his collaborator’s theories, Pauling went ahead and published his ideas on how antibodies work, hoping that further research could support his paper. Thus began a lengthy study of antigen-antibody binding in which Pauling and Campbell attempted to develop a complete theory. Along the way, Dan Campbell’s research at Caltech would become very important to the Institute as well as to Pauling.

In 1943 a Caltech research fellow named David Pressman agreed to join Campbell and Pauling in their study of immunology. Starting with work that had previously been published, Pressman, Pauling and Campbell refocused their studies to explain how antigens and antibodies bind, a change in focus from Campbell and Pauling’s earlier inquiries into how antibodies and antigens are formed. The decision to focus on previous research was made after Pauling had mistakenly announced that antibodies had successfully been synthesized at the Gates and Crellin Laboratories. As it turned out, attempts to create synthetic antibodies using Pauling’s proposed methods were completely unsuccessful. Pauling thus decided to start from scratch by developing a theory of antigen-antibody binding, which he would use to further investigate the chemistry of this interaction.

In July 1943, the three men published “The Nature of the Forces Between Antigen and Antibody and of the Precipitation Reaction,” appearing in the journal Physiological Reviews. The paper attempted to make more educated predictions about antigen-antibody binding.  In doing so, the article begins by referencing the concept of structural complementarity, which posits that antigen-antibody binding is driven by the close complementary physical shapes of the two molecules, which fit together like two adjoining pieces in a jigsaw puzzle. Commonly referred to as “the lock and key mechanism,” this idea was developed in the early 1930s, and served as Pauling and Campbell’s starting point in their initial investigations.

The 1943 study also drew from outside theories, such as the framework theory of precipitation, to suggest that antigen-antibody binding results in the formation of a precipitate; that is, that the two structures react to form an insoluble compound. Using these points as their foundation, the three researchers developed a new theory of antigen-antibody binding.

Pauling and Campbell, 1943.

Pauling and Campbell, 1943.

Campbell, Pressman and Pauling’s breakthrough came by way of their proposal that structural complementarity is an especially important feature for reactions that depend on Van der Waals forces. Van der Waals forces are weak forces of attraction that bind together molecules located in close proximity to one another. The Caltech researchers believed that the close complementary geometry of antibodies and antigens was significant enough to enable these molecules to fit together using the weak Van der Waals attraction as a binding force. In other words, the summation of Van der Waals forces present along the binding site of an antibody worked to bind it to its antigen, specifically because the shapes of antibodies and antigens complimented each other so closely. This theory explained much of what had been observed by immunologists across the discipline in multiple investigations of antigen-antibody reactions.

From here, the three researchers also asserted that two propositions placed forth in Pauling’s 1940 paper should still be considered for further study: the multivalence of antigen-antibody interactions and the probability of hydrogen bonds acting between the two molecules. The trio also concluded that the antigen-antibody mechanism would require at least two supplementary types of forces: Coulomb attraction and polar attraction.

Of the conclusions published by Campbell, Pressman and Pauling in 1943, the multivalence of antigen-antibody interactions and the three proposed forces (Van der Waals, Coulomb and polar) between the two molecules are still considered to be contributing factors to the functioning of the human immune system. With this publication, Campbell, Pauling and Pressman also showed that the immune system relies heavily on both structural and chemical features to carry out its processes.

The important conclusions derived from research conducted by Campbell, Pauling and others established Caltech as a leader in the field of immunology. Over the years that followed, Campbell and Pauling continued to develop their theory of antibody formation, which remained widely accepted until the 1950s. Even when the duo’s work began to be disproven by findings in the genetics field, the understanding of antigen-antibody interactions suggested by research done at Caltech remained undisputed.

Dan Campbell and Linus Pauling went on to publish more than twenty articles relating to immunology, exchanging ideas on the topic until the end of Pauling’s tenure at Caltech in the early 1960s. The attention that their work brought to the Gates and Crellin Laboratories at Caltech prompted the creation of a separate department, one that was entirely dedicated to immunochemistry. (The first of its kind on the west coast.)

For thirty years, Campbell headed Caltech’s immunochemical research and his fame as an immunologist grew to the point where, in 1972, he was named president of the American Association of Immunologists. Two years later, in 1974, Campbell passed away at the age of 67, the victim of a heart attack.  Over the course of his career, he published more than 200 papers as well as several books, and he served on editorial boards of four scientific journals related to immunology.

The Arrival of Dan Campbell at Caltech

Dan Campbell, ca. 1940s.

Dan Campbell, ca. 1940s.

[Part 1 of 2]

As a scientist, Linus Pauling is remembered by many for combining his expertise in chemistry with other fields. Often times Pauling would start off thinking about a problem from a chemical perspective and end up learning about a field entirely new to him, like cellular biology or medicine. Though this sort of cross-disciplinary work is more commonplace today (partly because of the example that Pauling provided), in the 1930s it was fairly rare for scientists to combine different fields of study. This given, pioneers of the cross-disciplinary approach often found it difficult to identify like-minded researchers with whom to collaborate. Fortunately for Pauling, a man with a very wide network, other researchers often found him.

After delivering a talk about hemoglobin in 1936, Pauling was pleasantly surprised to be consulted by Austrian medical researcher Karl Landsteiner. For many years, Landsteiner had been trying to understand how antibodies in the immune system work, and he believed that Pauling’s knowledge of medicine and chemistry could help him in his investigations. An antibody is a disease-fighting macromolecule that targets and rids the body of unwanted foreign substances, such as viruses and incompatible blood types. Landsteiner wanted to know how antibodies can target specific foreign substances with such precision. This encounter drew Pauling’s attention to the field of immunology, which would eventually become an important part of his research and would remain so for many years to come.

Pauling’s communications with Landsteiner spurred an interest in looking into the chemistry of antibodies and their substrates, antigens. At the time, however, most of Pauling’s focus was necessarily occupied with finishing up his previous program of grant-funded research on protein structures. Furthermore, Pauling was not an immunologist and the demands on his time were such that he could do little more than keep immunology in the back of his mind.

It wasn’t until 1939 that Landsteiner once again brought Pauling’s full attention back to antigens when he used Pauling’s theory of protein structure in a discussion about antibodies. Reading Landsteiner’s article sparked several ideas for Pauling which quickly led to his drafting a rudimentary theory of antibody chemistry. Six months later he found the perfect opportunity to test some these ideas.


Image extracted from a glass plate display, “Pictures of Antibodies,” prepared for the First International Poliomyelitis Conference, New York, 1948. The caption accompanying this image reads: “…[An] antibody-antigen framework which may precipitate from a solution or be taken up by phagocytic cells.”


In January 1940, immunologist Dan Campbell first visited Caltech on a fellowship. Campbell was an Ohio native who had been trained at Wabash College in Indiana and George Washington University in St. Louis, before receiving a doctoral degree from the University of Chicago, where he was subsequently hired as an assistant professor. During his tenure at Chicago, Pauling invited Campbell to spend a fellowship period at Caltech.  Campbell was only scantly familiar with the Institute, but was aware of the reputation of its chemistry department and accepted Pauling’s offer largely on this basis.

Due to his unfamiliarity with the institution, by the time of his arrival in Pasadena Campbell had still not yet identified a research project on which to collaborate. Pauling advised Campbell to consider different researchers before making his final decision on where and with whom he might work. In the end, after asking around, Campbell chose to collaborate with Pauling on his theory of immunology.

This was a fortuitous decision, for several reasons.  First, in addition to immunology, Campbell had a background in biophysics and chemistry, which made him a perfect candidate to test and develop Pauling’s antigen theory. More importantly, as Campbell began his initial investigations, it became apparent that Pauling’s ideas were flawed and that Pauling’s knowledge of chemistry alone would not be sufficient to make further progress in immunological research.


Campbell and Pauling, 1943.

Pauling had alleged that antibodies were similar to denatured proteins; that is, a protein that has lost its secondary and tertiary structures and has unfolded into an amino acid chain. Pauling’s theory anticipated that antibodies were an unfinished protein that required specific antigens in order to fold into the proper secondary and tertiary structures.

According to this model, antibodies would only form hydrogen bonds and thus would coil around chemically complementary antigens. As such, the theory explained how antibodies are able to bind unambiguously to their complementary molecules. However, Campbell’s results did not support all of Pauling’s ideas. Though his research showed that antibodies were in fact proteins, their physical structure before and after binding to antigens remained unclear.

Pauling’s lack of evidence for his theory of antibody structure and composition limited him to publishing only a single theoretical paper in which he explained his ideas about antibodies. In July 1940 the Journal of the American Chemical Society featured Pauling’s “A Theory of the Structure and Process of Formation of Antibodies.” The article received much attention and, despite the lack of evidence, was widely acclaimed, though it failed to provide a definitive explanation for antibody structure.

After the publication of the piece, Campbell once again tested Pauling’s theory, and this time his results were much more confusing, to say the least. Initially, it appeared that Campbell had succeeded in creating artificial antibodies by simply denaturing beef globulins (a protein found in blood) and later allowing them to refold around an antigen.

Word of these results greatly excited Pauling, who began to envision the mass production of antibodies using Campbell’s method. Reality turned out to be not so simple; when students and postdoctoral fellows tried to replicate Campbell’s experiment, they were unable to obtain the same results. Looking back now, it seems most likely that Campbell’s research assistants had misinterpreted the results of his experiment.

Pauling knew that he would need more time with Campbell to refine his theory, but that could only happen if Campbell’s position at Caltech was secured. In 1942 Pauling arranged for the Institute to offer Campbell an assistant professorship, which he accepted. By 1950 Campbell had become a full professor.

Combining immunology and chemistry proved to be a commendable approach for tackling many health concerns of the time. Likewise, Campbell’s presence was crucial to the development of Caltech’s immunochemistry department, which over a span of five years grew from a single office (Campbell’s) to a space occupying most of the third floor of Caltech’s Church Laboratory. Students and professors alike flocked to the growing department to discuss questions and engage in research on immunology, using chemistry as the basis of their approach. From the outset, both Pauling and Campbell benefited from one another’s expertise while colleagues at Caltech, and their partnership would continue to yield fruit for many years.

Rebecca Mertens, Resident Scholar

Rebecca Mertens

Rebecca Mertens

Rebecca Mertens of Bielefeld University, located in northwest Germany, is the latest visitor to complete a term as Resident Scholar in the Oregon State University Libraries Special Collections and Archives Research Center.  A Ph.D. candidate in the philosophy and history of science, Mertens spent a month stateside, visiting both the OSU Libraries as well as the Caltech Archives.

During her stay she braved both a major (and unusual) snow event in Corvallis as well as torrential rains in southern California.  Despite these obstacles, Mertens enjoyed a fruitful visit to the west coast as she pursued her research on Linus Pauling’s contributions to the lock-and-key model of biological specificity and the influence that this model imparted upon the sweep of modern biochemistry.

The conditions that awaited Mertens upon her arrival at OSU.

The conditions that awaited Mertens upon her arrival at OSU.

An outgrowth of his research on antibodies and antigens, Linus Pauling’s work on biological specificity comprised a major contribution to contemporary thinking on biochemical topics.  Pauling biographer Thomas Hager gives us this primer on what is meant by by the term, “biological specificity.”

Pauling demonstrated that the precise binding of antigen to antibody was accomplished not by typical chemical means – that is, through covalent or ionic bonds — but solely through shape. Antibodies recognized and bound to antigens because one fit the other, as a glove fits a hand. Their shapes were complementary. When the fit was tight, the surfaces of antibody and antigen came into very close contact, making possible the formation of many weak links that operated at close quarters and were considered relatively unimportant in traditional chemistry — van der Waals’ forces, hydrogen bonds, and so forth. To work, the fit had to be incredibly precise. Even a single atom out of place could significantly affect the binding.

In her Resident Scholar presentation, Mertens described the thrust of her research, which focuses on how one should interpret the contributions that Pauling made in this particular arena.

In the course of his research on antibodies, Linus Pauling postulated that the complementary structure of two molecules or two parts of a molecule determined the specificity of reactions in the living organism. However, the idea that molecular complementarity and biological specificity are deeply connected was already mentioned by Emil Fischer at the end of the 19th century. Thus, Pauling’s novel contribution was not the initial articulation of the model, but rather his emphasis on the importance of molecular complementarity for all biological phenomena.

Through examination of the Ava Helen and Linus Pauling Papers, as well as the institutional records held at Caltech, Mertens is pursuing the idea that “Pauling’s interdisciplinary reputation, his public presence and his engagement in the organization of scientific institutions led to the popularity of the lock-and-key model and to its standardization in the second half of the twentieth century.”  These forces of Pauling’s status and personality in turn made an impact on questions of “financial support, networking and science popularization within the administration of scientific projects.”

mertens-lecture

Beyond uncovering and detailing the history of Pauling’s role in the development of the lock-and-key model, Mertens is also using her research to “suggest an approach to the study of analogical models that considers social and political factors on successful model usage…[and] the formation and consolidation of model-based research programs.” Mertens returned to Germany with a large volume of content to sift through and absorb as she continues to develop her thinking on these issues.

Now entering its seventh year, the Resident Scholar Program at OSU Libraries provides research stipends of up to $2,500 to support work conducted in the Special Collections and Archives Research Center.  Applications for the 2014 class of scholars are being accepted now – the deadline for entry is April 30, 2014.  For more details, please see the program homepage.

The End of Artificial Antibodies

Illustration of bivalent antibodies attaching to complementary antigen molecules. Image extracted from a glass plate display, “Pictures of Antibodies,” prepared for the First International Poliomyelitis Conference, New York. The caption accompanying this image reads: “In vitro or in vivo bivalent antibodies may become attached to complementary portions of antigen molcules.”

[Part 3 of 3]

Though highly controversial, Linus Pauling’s claim that he had created artificial antibodies gave him a boost in funding. Many of his backers realized that if Pauling was correct he had just revolutionized modern medicine and they were just as eager as he was for his project to succeed. Since he now had more money, Pauling hoped he could expand his staff, though the war greatly prohibited this effort. Pauling lamented:

Unfortunately the amount of war work which is being done now here is so great that the usual seminars and informal discussions of science have decreased somewhat in number, but still a good bit of work in pure science is being carried on.

To help remedy this situation, Pauling corresponded frequently with William C. Boyd of Boston University’s School of Medicine, trying to persuade him to transfer to Caltech over the summer. Boyd refused, stating that he had too many other medical defense projects, though suggesting that “perhaps it will not be too late when the war is over, unless it goes on for 10 years or more, as some pessimistic writers predict.” Pauling also failed to hire two other well-known researchers, Henry F. Treffers, and A.M. Pappenheimer. Both declined because he was only able to offer a one year position. Indeed, from 1942-1943, Pauling actively tried to find staff for his lab, offering a $3,500 one-year position and draft deference. Despite this, most of the competent researchers he wanted were otherwise employed doing war work. He was able to hire Leland H. Pence in December of 1942, and in 1945 Frank Johnson visited from Princeton and worked at the lab in Caltech a bit. These individuals were, however, the exceptions, and Pauling was generally unsuccessful with his offers of employment.

Pauling was also facing other staff issues that were relics of the era. Among his group were two employees who were born and raised in the United States, but whose families were Japanese, Carol Ikeda and Miyoshi Ikawa. All too cognizant of the forthcoming policy of internment, Pauling began corresponding with Michael Heidelberg of Columbia University, hoping that he could temporarily trade employees as a method of getting Ikeda, at least, to a less hostile location. The plan did not work out, and Heidelberger ended one of his letters to Pauling bemoaning the fact that “…unfortunately a lot of wholly patriotic people are going to suffer.” Pauling was eventually able to get both Ikeda and Ikawa into graduate programs, though doing so took a substantial amount of work.


Dan Campbell and Linus Pauling, 1943.

Even though William Boyd had refused a job, he and Pauling continued to correspond frequently. Boyd was critical of Pauling’s theories on antibodies, warning his colleague that “preconceived notions evidently play a big role in the field [of immunochemistry.]” Boyd told Pauling to be careful with his research and his declarations, as he had often made arguments that he felt infallible only to have his colleagues inform him that they were unconvinced.

In August 1942, five months after issuing his controversial press release, Pauling finally published his research on artificial antibodies in an article titled “The Manufacture of Antibodies in vitro,” which appeared in the Journal of Experimental Medicine. As with the press release, Pauling’s paper was somewhat lacking in detail and many scientists found it hard to replicate his experiments. Those who did, such as Pauling’s valued colleague Karl Landsteiner, were unable to obtain the same results that Pauling had reported. Despite this, Pauling remained convinced that his research was valid and worth pursuing. However, Pauling’s collaborator Dan Campbell seemed to be the only one who could successfully produce the antibodies, and even those were weak and ineffective.

In early 1943 the Rockefeller Foundation assigned Frank Blair Hanson to assume some of the work that Warren Weaver had been conducting, and right away it was clear that Hanson was notably “less entranced than Weaver with Pauling’s work.” Despite the fact that he agreed to continue funding Pauling’s artificial antibody research, he was skeptical of its worth and began polling immunologists across the country to that end. They did not respond favorably – even Landsteiner believed that there was a less than 50% chance that Pauling had actually created artificial antibodies. As a result of these lackluster opinions, Hanson cut Pauling’s funding by half.

Pauling proceeded nonetheless, his enthusiasm still strong. His next move was to submit a patent application, “Process of Producing Antibodies.” The response that he received was not what he wanted to hear:

The claims are again rejected for lack of utility as no evidence has been presented to show that the antibodies alleged to be produced by the claimed have any utility at all… The claims are rejected as being too broad, functional, and indefinite…the claims are rejected for lack of invention…all the claims are rejected.

Pauling was disheartened by this response but still confident that he could salvage the project. However, shortly afterward he received a letter from the Rockefeller Foundation informing him that they did not approve of researchers using their funds to apply for medical patents. Because of the letter, and the general ineffectiveness of the research, Pauling opted not to pursue his patent application any further.

Illustration of the antibody-antigen framework. The caption accompanying this image reads: “…[An] antibody-antigen framework which may precipitate from a solution or be taken up by phagocytic cells.”

As 1943 progressed, Pauling continued to piece together a better understanding of how antibodies adhere to antigens. Later that year, he and Campbell published a paper in Physiological Review which further elaborated on their idea that shape was the primary determinant of antibody functions. They wrote that while many of their colleagues at other institutions felt that antibody formation adhered to a “lattice theory,” they did not, because their research showed that the structures created by antibody/antigen precipitation were not regular enough. Instead, they coined the phrase “framework theory” to describe their idea.

By the winter of 1943-1944, Pauling had at last concluded that the artificial antibody research was going nowhere. This was a difficult admission for Pauling because, despite the fact that progress in the research was still slow, he was convinced that he could make it work if given more time. Unfortunately for him, the war effort demanded quicker results and actively prohibited greater focus on antibodies. In the end, he decided to abandon the artificial antibodies research. Pauling never retracted his support for the work, though many years later Dan Campbell admitted that a laboratory technician had “shaded” the results to fit what he thought Pauling wanted to see.


The failure of the artificial antibodies project allowed Pauling to move on to more productive lines of research. He continued to build his ideas on how exactly antibodies function, and by 1945 he was able to prove that shape was indeed what caused antibodies to adhere to antigens. In Pauling’s description, the antibodies would fit to the antigens like a glove, at which point they adhered, not due to orthodox chemical bonds, but because of another weak, poorly understood force.

The force that causes antibodies to bind with antigens is called the van der Waal’s force. It is a very weak, almost imperceptible subatomic bond between two molecules existing in extremely close proximity. Historically, due to their weakness, van der Waal’s forces had been ignored as viable components of biochemical reactions. However, Pauling was able to show that the extremely tight fit between antibodies and antigens exposed a large surface area across which the van der Waal’s force could become a factor. The fit had to be precise, as even one or two atoms being out of place would effectively break the hold that the van der Waal’s force put into place. This concept, known variously as molecular complementarity or biological specificity, cast a great deal of light on a central mystery of molecular biology. With it, Pauling was additionally able to confirm his earlier hypothesis that antibodies are bivalent.

As World War II drew to a close, Pauling shifted his focus away from antibodies and back towards a more general study of the shape, creation, and function of proteins. Pauling’s focus on proteins was long lived, stretching at least twenty-five years from 1933-1958. His foray into antibodies was notably shorter, a nine-year interlude from 1936-1945. Yet in this time, he had managed to dramatically impact immunochemistry with his discovery that antibodies are bivalent and his insights into how they work. These accomplishments are made even more impressive when considering that Pauling was neither an immunochemist nor an immunobiologist by trade. As was so often the case during his life, he threw himself into the challenge with characteristic enthusiasm, and managed to make major contributions to the field.

The Tantalizing Prospect of Artificial Antibodies

Linus Pauling and Dan Campbell, 1943.

[Part 2 of 3]

By late 1939, Linus Pauling had thrown himself wholeheartedly into the study of antibodies, specifically how they work and how they are made. He’d already developed a few memorable and unique hypotheses, though by 1940 they were still yet to be proven correct or otherwise.

In January 1940, a researcher named Dan Campbell arrived at Caltech with the intent of working on problems in immunochemistry. He and Pauling began collaborating, and ended up co-authoring several papers together. Later in 1940, after a first, erroneous paper, Pauling published another in which he claimed:

all antibody molecules contain the same polypeptide chains as normal globulin, and differ from normal globulin only in the configuration of the chain; that is, in the way that the chain is coiled in the molecule.

In other words, shape was what determined the effect of antibodies. Pauling acknowledged the potential for flaws lying within his hypothesis, but adopted it because of his inability to otherwise “formulate a reasonable mechanism whereby the order of amino-acids residues would be determined by the antigen.”

Nobody knew about the genetic basis of amino acids at the time. As a result, while Pauling’s hypothesis on how antibodies worked eventually turned out to be correct, his coupled hypothesis on how they were formed was still wrong. Regardless, Pauling’s theory “ruled the roost amongst immunochemists” for almost 20 years. It wasn’t even until 1949 that people began to seriously question Pauling’s model, and when that finally happened, it was because the model failed to account for the development of immunity to antibodies.

In July a colleague at Caltech, Max Delbrück – a German researcher who had arrived in the US on a study abroad trip and never went home – showed Pauling a paper written by Pascual Jordan, another German scientist. Jordan claimed that identical molecules were attracted to each other, and opposite molecules repelled from one another, because of quantum mechanical resonance. Pauling declared the idea “baloney,” and decided to write a paper debunking Jordan’s theory. He asked Delbrück to co-author it, which he did with hesitation, as the young scientist was nervous about signing his name to a paper attacking the theories of the famous and well-respected Jordan. The duo finished the paper and published it in Science, but unfortunately for them, the work went largely unnoticed.


Since the start of the Battle of Britain in 1940, Pauling had been doing some side research on explosives, propellants, and other “war work” in anticipation of United States involvement in the war raging across Europe and the Asia. In 1941, U.S. engagement finally came about with the attack on Pearl Harbor. Pauling and his laboratory at Caltech switched their main priority to war work, as did just about every other research laboratory in the country. As part of this realignment of efforts, Pauling received a grant to begin a research project with the goal of creating gelatin-based blood plasma artificial antibodies, to be used for military medical purposes.

Pauling hypothesized that if a generic protein, such as beef globulin, was very carefully denatured, then placed in the presence of an antigen and revitalized, it would grow and mold itself to the antigen. If this worked, medical technicians would someday be able to create antibodies “made to order.” The potential import of the idea was clear: “the end result of a million years of evolution” would become available “by the quart.” Successful implementation would represent a massive breakthrough in medical technology, and Pauling was convinced that he was just the man to do it. He picked Dan Campbell and David Pressman to be his primary assistants and the group got to work.

The process was slow and complicated; it involved a mixture of 1-2% beef globulin, dissolved in 0.9% sodium chloride, then very slowly heated to 57˚F. Once at that temperature, alkali would be carefully added to bring the mixture to a pH of 11, before very delicately and slowly returning the mixture to pH 0. To further complicate issues, the process produced a large amount of nitrogen, which had to be removed, but the process of removing the nitrogen created carbon monoxide and other “undesirable substances.” The Pauling group tested their antibodies on rabbits, mice, and guinea pigs predominantly, as their labs lacked the space for more extensive animal research.

In March 1942, instead of submitting his ideas to a peer-reviewed scholarly journal, Pauling issued a press release to Science News, claiming that his lab had developed artificial antibodies.

Text of Pauling's artificial antibodies press release, March 1942.

Text of Pauling’s artificial antibodies press release, March 1942.

The scientific community was vexed by this highly unusual action. Numerous scientists questioned the strength of the data, saying that the reports released by Pauling and Campbell were vague and weak. Pauling asserted that while the data they had gathered was not definitive, it was strong enough to make the statement that artificial antibodies had been produced. Pauling said that his artificial antibodies, though weak, were still real, and that he was going to continue improving on them. Specifically, he claimed that the antibodies had been used to prevent mice from getting pneumonia.

Pauling’s reputation helped to coax certain colleagues in the direction of his arguments. And after some skillful debating, the Rockefeller Foundation offered $31,000 to continue the research, while the federal Office of Science Research and Development offered $20,000.

Despite the skepticism of his peers, Pauling was enthusiastic and eager to continue research on this line of work. He had the funding and the desire, and he was convinced it was only a matter of time before he could start mass-producing artificial antibodies. However, he was about to run into some issues outside of his laboratories.

Thinking about the Creation of Antibodies

Drawings of the interaction between an antibody and azoprotein by Linus Pauling. 1940s.

[Part 1 of 3]

During a period of about a decade, beginning in 1936, my principal research effort was an attack on the problem of the nature of life, which was, I think, successful, in that the experimental studies carried out by my students and me provided very strong evidence that the astonishing specificity characteristics of living organisms…is the result of a special interaction between molecules…

-Linus Pauling, 1991

As one would expect, the Great Depression made it extremely difficult to acquire funding for scientific research. Luckily for Linus Pauling, he had on his side the patronage of the Rockefeller Foundation, and a close friendship with Warren Weaver, the head of the foundation’s Physical Sciences Division. Weaver wanted Pauling to get more involved in biological research, specifically protein research. Pauling was hesitant, but Weaver controlled the funding and convinced Pauling to move forward.

Alfred Mirsky

By 1933 Pauling was heavily involved in research on proteins, specifically their shape and function. In 1935 he began working with Alfred Mirsky, a Rockefeller Foundation scientist, with whom Pauling concluded that all proteins are structured as chains, and that the shape of a given protein determines its function and behavior.

A large component of Pauling and Mirsky’s research was on protein denaturation, effectively the breakdown or death of proteins. It was known that modest heating, mild acids, milk alkalis, or agitation, such as beating eggs with a fork, all serve to denature a protein. However, Mirsky discovered that proteins that were slowly denatured at lower temperatures could be resuscitated and the process reversed.

Karl Landsteiner

In the Spring of 1936, Pauling began another collaboration, this time with Karl Landsteiner, an Austrian scientist who won a Nobel prize for discovering and developing the field of blood typing. Landsteiner invented the ABO system, and uncovered methods for making blood transfusions safe. In this research Landsteiner observed that, in instances where the wrong blood type is used in a transfusion, antibodies attacked the transfused blood. Pauling was intrigued by Landsteiner’s work, and began reading about antibodies; he was interested and puzzled by what he found. While the scientific community knew that antibodies worked, how exactly they worked and how exactly they were formed were still unknown.

At the time, there were four main schools of thought regarding the creation of antibodies: the Antigen-Incorporation theory, the Side-Chain theory, the Instruction theory, and the Selection theory.

The Antigen-Incorporation theory, originally proposed by Hans Buchner in 1893, proposed that antibodies were actually the byproduct of antigens “splintering” in the human body and becoming incorporated into it. Despite the fact that this theory had been largely disproven at the time, it was proposed again by E. Hertzfeld and R. Klinger in 1918, by W.H. Manwaring in 1926, by Locke, Main, and Hirsch also in 1926, and finally once more by Gustave Ramon in 1930.

The Side-Chain theory was posited by the famous Paul Ehrlich in 1897, who argued that the body’s immunological reaction to antigens was “only a repetition of the processes of normal metabolism.” Ehrlich thought that cells would digest certain antigens in the same way that they digested nutrients. After repeated assimilations, or too large of an assimilation, the cells would overcompensate and release antibodies. His theory included a number of issues that the scientific community could not solve at the time, and it took over sixty years for the model to be improved upon.

The Instruction theory states that the body uses antigens as a template, then manufactures antibodies to specifically combat the antigen that the antibody is based off of. Pauling eventually belonged to this school of thought, as did Landsteiner, Michael Heidelberger, Felix Haurowitz, and Jerome Alexander. This group was far from unified however; the only point on which adherents to this school agreed was that antigens acted as templates. How antibodies worked, and how they were produced, was still a highly contentious question.

The final theory was the Selection theory, which was in concept almost identical to Ehrlich’s Side-Chain theory, except that its explanations were based on more modern mechanisms. Instead of general metabolic processes, quantum mechanical forces were proposed to be the cause of the attraction between antigens and antibodies. This school of thought became more popular near the end of World War II and in the post-war era.

Pauling described Antibodies as “fantastically precise little weapons,” and found it fascinating that they could identify and attack invading molecules that were different from safe molecules by only a few atoms. Antibodies are made of pure protein, are remarkably similar to one another, are relatively enormous, and also attack vastly different types of molecules.

Pauling and Landsteiner were especially vexed by how antibodies could target varied molecules so precisely when they were so similar. Pauling proceeded to read Landsteiner’s book on antibodies, and began to wonder if shape affected antibodies as much as it affected regular proteins. Landsteiner had arrived at a similar conclusion, and in 1939 published a note in Science suggesting that shape was what determined the effect of antibodies.

Diagram included in Pauling’s article, “A theory of the structure and process of formation of antibodies.” 1940.

Pauling expanded upon this idea, and in 1940 published a paper in which he hypothesized that antibodies were built as chains of non-specific proteins which collided with antigens, then compressed and shaped themselves around the antigen, “like wet clay pressed against a coin.” The paper created quite a stir, and generated a lot of support for the notion of using chemistry to solve biological questions. Unfortunately for Pauling, it later turned out that his hypothesis was deeply flawed.

Another argument developed in the 1940 paper was that antibodies are bivalent – that is, they have two sites which can bind to antigens. In addition to being bivalent, Pauling hypothesized that each of the “arms” of an antibody could latch onto different kinds of antigens. While Pauling was incorrect on the latter part – antibodies can only grab onto one type of antigen – he was correct that they are bivalent.

Pauling had gotten off to a strong and noticeable start in the field of immunology. Whether correct or incorrect, he was making progress towards a greater understanding of how the body protects itself. As the clouds of war began to reach across the Atlantic and Pacific towards the United States, Pauling’s new and growing knowledge was going to be put to the test.

Linus Pauling and the Structure of Proteins: A Documentary History

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Today is Linus Pauling’s birthday – he would have been 112 years old.  Every year on February 28th we try to do something special and this time around we’re pleased to announce a project about which we’re all very excited: the sixth in our series of Pauling documentary history websites.

Launched today, Linus Pauling and the Structure of Proteins is the both latest in the documentary history series and our first since 2010’s The Scientific War Work of Linus C. Pauling. (we’ve been a little busy these past few years)  Like Pauling’s program of proteins research, the new website is sprawling and multi-faceted.  It features well over 200 letters and manuscripts, as well as the usual array of photographs, papers, audio and video that users of our sites have come to expect.  A total of more than 400 primary source materials illustrate and provide depth to the site’s 45-page Narrative, which was written by Pauling biographer Thomas Hager.

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Warren Weaver, 1967.

That narrative tells a remarkable story that was central to many of the twentieth century’s great breakthroughs in molecular biology.  Readers will, for example, learn much of Pauling’s many interactions with Warren Weaver and the Rockefeller Foundation, the organization whose interest in the “science of life” helped prompt Pauling away from his early successes on the structure of crystals in favor of investigations into biological topics.

So too will users learn about Pauling’s sometimes caustic confrontations with Dorothy Wrinch, whose cyclol theory of protein structure was a source of intense objection for Pauling and his colleague, Carl Niemann.  Speaking of colleagues, the website also delves into the fruitful collaboration enjoyed between Pauling and his Caltech co-worker, Robert Corey.  The controversy surrounding Pauling’s interactions with another associate, Herman Branson, are also explored on the proteins website.

Linus Pauling shaking hands with Peter Lehman in front of two models of the alpha-helix. 1950s.

Linus Pauling shaking hands with Peter Lehman in front of two models of the alpha-helix. 1950s.

Much is known about Pauling’s famously lost “race for DNA,” contested with Jim Watson, Francis Crick and a handful of others in the UK.  Less storied is the long running competition between Pauling’s laboratory and an array of British proteins researchers, waged several years before Watson and Crick’s breakthrough.  That triumph, the double helix, was inspired by Pauling’s alpha helix, discovered one day when Linus lay sick in bed, bored and restless as he fought off a cold. (This was before the vitamin C days, of course.)

Illustration of the antibody-antigen framework, 1948.

Illustration of the antibody-antigen framework, 1948.

Many more discoveries lie in waiting for those interested in the history of molecular biology: the invention of the ultracentrifuge by The Svedberg; Pauling’s long dalliance with a theory of antibodies; his hugely important concept of biological specificity; and the contested notion of coiled-coils, an episode that once again pit Pauling versus Francis Crick.

Linus Pauling and the Structure of Proteins constitutes a major addition to the Pauling canon. It is an enormously rich resource that will suit the needs of many types of researchers, students and educators. It is, in short, a fitting birthday present for history’s only recipient of two unshared Nobel Prizes.

Happy birthday, Dr. Pauling!

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