A year before his official retirement from Columbia University in 1955, Heidelberger joined the Institute of Microbiology at Rutgers University at the invitation of its founder, Selman Waksman. Waksman had endowed the institute with patent proceeds from his discovery of streptomycin, and had established within it a laboratory of immunochemistry and immunology under Otto J. Plescia, with whom Heidelberger had studied complement fixation at Columbia University's College of Physicians and Surgeons during the 1940s. Personal connections reached further still, as Waksman's son, Byron, had studied components of complement in Heidelberger's laboratory in 1950. The field of immunochemistry in the mid-1950s was still small, many of its practitioners having been personally trained by Heidelberger and his students.
Together with Plescia, Heidelberger instituted a graduate course in immunochemistry at the institute, an expansion of the lectures on immunology and immunochemistry he had regularly given to medical students at Columbia. The half-year course, offered every other year, included laboratory work during which the one dozen students immunized rabbits with egg albumin, a water-soluble glycoprotein, which they had isolated from newly-laid eggs and then crystallized. They analyzed the precipitate obtained by adding recrystallized albumin, along with a second protein, egg globulin, to the antialbumin serum produced in the rabbits, using Heidelberger's quantitative precipitin method to measure, isolate, and purify anti-albumin antibodies. In the course of this exercise the students learned just how difficult it is to purify a protein: sometimes their results contained as much antiglobulin as antialbumin. At the institute Heidelberger also sponsored his last graduate student, Sergio Estrada-Parra of the Polytechnic Institute of Mexico City, who continued the work of Harold Markowitz on type XVIII pneumoccoal polysaccharide, for which Estrada-Parra was able to identify two alternative chemical structures.
Having lived for five decades on New York's Upper West Side and worked at institutions a short subway ride away, commuting to Rutgers was a new challenge for Heidelberger and his 1939, four cylinder Oldsmobile. During the first winter he often had to stop along the way and scrape ice from the windshield because he could not get a heater for his obsolete car. After he and his second wife, Charlotte Rosen, were married in 1956, they rented an apartment near campus for their stays in New Brunswick.
Heidelberger continued to travel widely during the 1950s, both to scientific meetings and on behalf of the United States Association for the United Nations, an organization that helped promote the efforts of the world body to achieve peace through multilateral negotiations and agreements between nations. He attended the first Asian meeting of the World Federation of United Nations Associations, in Bangkok in 1955. His resolution in favor of an international peace conference, edited and promoted by Eleanor Roosevelt, was not adopted. More successful were the six lectures he delivered en route at the medical school of the University of Tokyo at the invitation of Tomio Ogata. They were recorded and published as a book, Lectures in Immunochemistry (1956).
In 1964, Heidelberger returned to a medical school setting, with its widely varying clinical and basic research interests, when he moved to the Department of Pathology in the New York University Medical School. There, his colleagues in immunology included Zoltan Ovary, an expert on anaphylactic hypersensitivity, and Baruj Benacerraf, a pioneer in the study of cellular immunity and future Nobel Laureate. Drawing on his extensive stock of antisera and polysaccharides, Heidelberger was able to continue his research on a shoestring budget, foregoing by the 1970s even his modest National Science Foundation grant to the benefit of younger scientists.
Some of Heidelberger's research at NYU during the 1960s was designed to localize immunologically reactive groupings of complement on the surface of proteins and antiproteins. However, for most of the years after 1954, he worked largely on immunological cross-reactions between different species of microorganisms such as Klebsiella and Escherichia coli bacteria, and various antipneumoccoal sera. From these cross-reactions much could be learned about the chemistry of polysaccharide antigens. In particular, Heidelberger was able to construct from cross-reactions hypotheses regarding the molecular structures of polysaccharide substances, in particular the placement of their sugars, a key factor in understanding their antigenic properties and thus their disease-causing potential. The main thrust of these experiments, then, was to correlate chemical structure and immunological specificity, the same overarching objective he had pursued ever since his and Oswald Avery's pioneering studies of the polysaccharide antigens of pneumococcus during the 1920s.
For example, Heidelberger helped discover that while many thread-like polysaccharide antigens have either cellobiuronic or aldobiuronic acids (both disaccharides made up of two monosaccharides) as the main components of their repeating chemical units, others, such as type IV and type XXVII pneumococcus polysaccharide as well as a type of Klebsiella capsular polysaccharide (K81), contained pyruvic acid. His finding that pyruvic acid, an essential element in the metabolism of most living cells, was also of immunologic importance surprised Heidelberger, but became irrefutable when he and William F. Dudman showed that its removal caused a profound change in the immunologic specificity of the polysaccharide antigens. Namely, removal prompted a drastic reduction in immune precipitation with homologous antisera (antisera to the specific types of Klebsiella and pneumococcal antigens under examination) on the one hand, and on the other hand produced cross-reactivity with several non-homologous types of antipneumococcal sera (antisera formed against a different type of pneumococcal antigen with which the first antigens, here Klebsiella K81 and type IV and type XXVII pneumococcal polysaccharide, share closely related surface sites called antigenic determinants). Both results were to be expected if pyruvic acid was in fact immunodominant, the former because the substance (the acid) that controlled the chemical reaction between antigen and homologous antisera had been eliminated, the latter because its removal freed two sugar hydroxyls that had been previously blocked by the pyruvic acid, and which could now react with chemical groups on the surface of non-homologous antisera.
Similar cross precipitations were later observed between antipneumococcal sera and many other of the approximately eighty types of Klebsiella capsular polysaccharides (over twenty of them precipitated by one antiserum alone, anti-PnII), another surprising finding given the profound evolutionary differences between pneumoccocus and Klebsiella. A scientist at the University of Rostock in East Germany had purified the Klebsiella polysaccharides and sent it to Heidelberger. In fact, colleagues continued to send him materials for analysis almost until his death at 103.
From comparisons of these cross reactions, Heidelberger could once again deduce, for example, that certain reacting Klebsiella types had D-glucuronic acid or L-rhamnosyl residues as lateral, non-reducing end groups (meaning that they did not take up hydrogen or electrons). Heidelberger concluded that many types of Klebsiella, with different kinds and combinations of constituent antigens, could be grouped into chemical types according to the nature and position of a sugar that determined each type's immunological reactivity. Similar chemical classifications based on the position of immunodominant sugars were undertaken by other scientists for additional microorganisms, such as by Heidelberger's friend Otto Westphal for Salmonella. By way of summing up over four decades of research on cross reactions, Heidelberger at the time of his death in 1991 was preparing a lengthy manuscript, "Cross reactions of 56 bacterial polysaccharides with 24 antipneumococcal, three antisalmonella, one antiklebsiella and antimyoplasma sera."