Figure 1. Sir Gustav Nossal. Sir Gustav Nossal has been a member of the Editorial
Board of Immunology Today/Trends in Immunology since the journals inception,
25 years ago.
Review TRENDS in Immunology Vol.25 No.12 December 2004of ideas!activities, reputations and influences can spring.Gus, like one of his very early mentors Sir Macfarlane
Burnet, has subscribed to the view that basic science isprimarily about ideas. Creating the right environment forscientists, to enable ideas to emerge, where vigorousdebate flourishes and where hypotheses are put to thetest and bureaucracy minimized is what scientific leader-ship is about. One of his best-known quotes is . theglittering prizes of success come to those who combineteamwork with high technology in a bubbling cauldron
Despite the primacy of ideas in basic scientific research,Gus interests in translating and implementing (evencommercializing) the products, processes and services ofresearch have strengthened over time. He has involvedhimself deeply at the global level in both development ofnew vaccines and increasing population access to those inexistence, particularly childhood vaccines. A completepicture of Gus Nossals contribution to immunology willtherefore comprise laboratory-based scientific researchoutputs, efforts to translate discoveries to products ofscientists credibility and from which the other interests,
ceptual and technological currency. This is the basis of aHistorical review: Simmunologist and mGraham F. Mitchell1 and Michael F. Good1Foursight Associates Pty Ltd, Level 2, 164 Flinders Lane, Melbo2Queensland Institute for Medical Research, 300 Herston Road,
Sir Gustav Nossal, universally known as Gus Nossal, isAustralias best known medical research scientist andadvocate, who over the past five decades has influencedthe course of immunology. His research interests in thisfield commenced during PhD studies, and his accom-plishments centre around the cellular events of antibodyproduction. His scientific achievements are matched bythe energy he has devoted over his entire career topublic health; science in society; political, philanthropicand community support for science; global access toexisting vaccines and development of new vaccines; andthe personal and scientific development of colleaguesand young scientists. His contribution and philosophy ofscience, together with political influence, capacity toinspire and an extraordinarily engaging personality,place Gus Nossal in the top bracket of biomedicalscientists of the second half of the 20th century.
For almost 50 years, and certainly over the 25-year historyof Trends in Immunology and Immunology Today, GusNossal (Figure 1) has consistently and proudly describedhimself in a multitude of forums as an immunologist andoccasionally, as appropriate, an Australian immunolo-gist. His professional life as such is a reflection of a deepconviction that, despite encyclopaedic scientific knowledgeand infinite capabilities, genuine interests in all branchesof medicine, major ongoing societal contributions andserious political influence, above all else, the essence of amedical researcher is deep expertise in a particulardefined research area. In one discipline, specialty orresearch endeavour, depth rather than breadth is para-mount and, by definition, must be accompanied by con-Gustav Nossal ore
e, Victoria 3000, Australiaston, Queensland 4006, Australiacareers of a multitude of immunologists and medicalresearchers more broadly.
Corresponding author: Graham F. Mitchell (email@example.com).Available online 30 September 2004
www.sciencedirect.com 1471-4906/$ - see front matter Q 2004 Elsevier Ltd. All rights reserved. doi:10.1016/j.it.2004.09.007public health and commercial value and impacts on the
The B-cell immunologistLike many of us, Gus is best known among immunologistsfor his early research; unlike many of us, he has stayedpretty much with a line of research that commencedduring his PhD studies: the cellular events of antibodyformation, extending to immunological memory andimmunological tolerance. Moreover, the vast bulk of hisresearch work was performed at one institution, namelyThe Walter and Eliza Hall Institute of Medical Research(WEHI) in Melbourne. Being concerned with the inductiveevents of immune responsiveness and the detailedcharacteristics of antibody-producing cells and theirprecursor lymphocytes, he is primarily a B-cell immunol-ogist. A particular source of pride has been continuous(19621996) National Institutes of Health (USA) grantsupport (AI-03958) for studies on antibody production atthe single-cell level.
The early yearsAs the focus of this article is the contribution of Sir GustavNossal to the field of immunology in the years since hiscontribution to the first issue of Immunology Today (i.e. the second half of his immunological life), we willsimply headline some of his early research contributions
lymphocytes (B cells) from non-immunized animals isolated through
Review TRENDS in Immunology Vol.25 No.12 December 2004666antigen-binding fractionation; Development of robust micromanipulation and other cell-separation techniques to visualize and study single cells (later,exploitation of carefully optimized limiting-dilution in vitro clonalanalyses); Demonstration of a switch in isotype from multivalent IgM todivalent IgG in individual antibody-producing cells without changein specificity for antigen; Contributions to immune response maturation from early IgM toIgG antibodies with subsequent antibody affinity increase; Demonstration of follicular localization of (radiolabelled) antigenin secondary lymphoid tissues (i.e. lymph nodes and spleen) androle of antibody; Demonstration of persistence of (radiolabelled) antigen on follicu-lar dendritic cells; Insights into the development of germinal centres from primarylymphoid follicles; Highlighting the importance of antigen characteristics (e.g. valency,solubility), antigen dosage (i.e. low- and high-dose tolerance) andage of exposure in tolerance induction; Demonstration through chromosome analyses of micromanipu-lated antibody-secreting cells that, in T cellB cell collaboration, only(Box 1), many through collaboration, which, incidentally,was always acknowledged, often lavishly.
Among the numerous research contributions anddiscoveries of the early years outlined in Box 1, two havehad a major impact. First, the use of micromanipulationtechniques to study single antibody-producing cells, withthe data strongly supporting, if not proving, the clonalselection theory of Burnet. Second, the equally pain-staking pioneering work with Gordon Ada and other
Box 1. Research contributions (late 1950s to mid-1970s)
The one cellone antibody rule; more precisely, the one celloneantibody specificity rule; Absence of (radiolabelled) antigen in antibody-producing cells; Validation of the clonal selection theory of antibody formation,including the demonstration of unique specificity of individualB cells produce antibody.
www.sciencedirect.comcollaborators that demonstrated the trapping of antigenon follicular dendritic cells of lymphoid follicles andgerminal centres i.e. follicular antigen localization.
More recent researchIn the decade before his retirement from WEHI in 1996,his research discoveries, again through collaboration(primarily with PhD students, postdoctoral fellows andsenior technical staff), were predominantly in two areas the mechanisms of B-cell tolerance and identification ofkey decision points for tolerance induction over thedifferentiation history of nave and sensitized B cells. Wewill now outline the essential features of this morerecent work.
If the mechanisms of immunological tolerance at theB-cell level are viewed as a spectrum, with the concep-tually straight forward and well-grounded clonal deletionof Burnet at one end, then the more complex, andseemingly risky, clonal anergy of Nossal is at the otherend of the spectrum. Inhibition of antibody production,and thus control of autoantibodies in particular, appar-ently comes in several guises: yet another example of thebody not investing in only one mechanism to achievecrucial biological, and thus evolutionarily important,outcomes.
The work of Nossal et al.  on clonal anergy that beganin the late 1980s demonstrated that self-destruction wasnot the only choice available to B cells when inhibition ofantibody production is the desired outcome. Rather than aselective purging of the repertoire in a situation of B-celltolerance, antibody receptor-bearing B cells could still bedemonstrated. Clonal anergy, a selective silencing (ratherthan purging) in the repertoire, seemed to be an optionfor B cells, particularly at certain time points in theprocess of B-cell development and differentiation in bonemarrow and lymphoid tissues. Thus, B cells could beparalysed and, although still demonstrating specificityfor antigen, were prevented from progressing to high-levelantibody secretion.
The above experiments were performed using foreignantigens known immunogens (and tolerogens). In thecase of autoantigens and maintenance of self-tolerance, itwould seem risky to foster the presence of reactive yetclonally anergic B cells that might have some antigen-presentation capabilities for T cells. Just as B-celltolerance apparently involves more than the purging ofB cells, mechanisms of T-cell tolerance appear to bemultiple, and it is feasible that anergic B cells might beinvolved in maintaining sensitized regulatory T cellsin immunological tolerance. Alternatively, it is inter-esting to speculate whether the elimination of autoreac-tive B cells with the capacity to bind and presentautoantigens, at least to sensitized T cells, provides arationale for the use of particular B cell-depletingmonoclonal antibodies in the treatment of autoimmunedisease (D. Tarlinton, pers. commun.).
Across immunology, the impact factor of clonal anergyhas been modest. Despite the considerable uncertaintyaround the extent, nature and physiological importance of
this phenomenon, the notion that B cells pass throughperiods of heightened antigen-dependent susceptibility to
negative signalling has clearly stood the test of time. It isan immunological paradigm. There are key decision pointsin B-cell differentiation where the consequences of antigenencounter are weighted towards either negative orpositive signals. Additional considerations will clearly bethe presence of T-cell cytokine products, danger signalsfrom microorganisms and Toll-like receptor-bearing cells,and other aspects of the microenvironment local to
1950s, Gus has been a devotee of quantitation, as
Review TRENDS in Immunology Vol.25 No.12 December 2004 667quantitative analysis is a key ingredient of analyticalrigour. Like his scientific contemporaries and SydneyUniversity alumni colleagues Don Metcalf and Jaq Miller,and in keeping with the approach championed by NielsJerne and Mel Cohn, Gus recognized early on that the bestway to address the uncertainties of biological phenomena(and certainly the sometimes problematic assays ofimmune responsiveness, despite being at least based onspecificity and thus having an inbuilt control) was to getthe numbers and do the stats. Little is black and white inbiology, and the area of grey is best accommodatedthrough rigorous quantitation, in this case over the timecourse of an immune response (e.g. numbers of antibody-producing cells and numbers of their precursor Blymphocytes). Of course, it was left to Jerne et al.  toprovide the ultimate means to compare in vivo immuneresponses at the cellular level at that time the plaque-forming cell assay for antibody production to sheeperythrocytes. How, for years, we longed for a comparable
Box 2. Germinal centres
The demonstration some years ago that germinal centres are sites ofhypermutation in Ig V genes brought Nossal and Adas follicularlocalization studies  back into focus. Antigen localization ismarkedly influenced by existing antibody levels, and the overallavidity of any new B-cell receptor would need to be such that itcompetes for antigen with the range of existing antibodies,presumably anchoring that antigen on the follicular dendritic cell.antigen-binding B cells (which, at the population level,will also display a spectrum of avidity for antigen). Thisline of work extended to Klinmans second window ofsusceptibility to tolerance induction , this time ingerminal centres (the sites of hypermutation in B-cell Vgenes) (Box 2), perhaps through apoptotic mechanisms .
Virtually all of the work outlined above was performedin non-transgenic systems, with use of labour-intensiveflow cytometry and fluorescence-activated cell sorting andsingle-cell cloning analyses. Interestingly, confirmatorydata on clonal anergy have come from studies intransgenic mice . Gus and co-workers have investedheavily in developing and adapting new technology butmost obviously, in devoting the necessary time to prelimin-ary work on optimizing techniques and reagents that aregoing to be the linchpin of future research. Interpretationof results must not be clouded by technological deficienciesand uncertainties; emphasis must be on what doeshappen versus what can happen.
The numerate immunologistAs evidenced in his early publications at WEHI in the lateThere would thus be selection of B cells bearing mutated V genes ofprogressively higher binding site affinity.
www.sciencedirect.comassay of T-cell numbers engaged over time in an in vivoimmune response: the significant impedance being thatwe did not have a clue what they produced.
It is fascinating to look back at the early publicationsfrom Gus on the one cellone antibody demonstrationsand the emphasis on the arithmetic; that is, the numberof micromanipulated antibody secretors isolated fromanimals immunized with two antigens and comparingthe number producing antibody specificity 1 versusantibody specificity 2 versus possible double producers,and making comparisons with the precise numbersobtained by others using different techniques . More-over, quantitation was crucial in his determination of thecellular events of memory and tolerance, and the esti-mation of the number of antibody molecules produced by aplasma cell (10 000 per second) was another example.
This ease and familiarity with numbers finds itsexpression these days in the speed with which Gus canzero in on the key features of a companys financialstatements, a skill apparently learnt as a young boy fromhis businessman father on Sunday nights, while pouringover the book of accounts! Although not particularlyrelated to numeracy, another key element of Gusapproach to science that he shares with Miller andMetcalf is getting citations right, and their students allacquired this message at an early stage in their studies.The major purpose of at least Chapter 1 in a PhD thesis isto understand the background science in enough detail tobe able accurately to ascribe a relevant finding to apa...