Chapter I1 The Structure of Rheumatoid Factors Chairman: HENRYG . KUNKEL, M.D. DR.KUNKEL: The topic of the structure of rheumatoid factors revolves primarily around the problem of whether or not the rheumatoid factors as seen in rheumatoid arthritis sera are antibodies; the question of the structure of these “antibodies” is not one we should get into here. There follows a list of some of the properties of rheumatoid factors which favor the hypothesis that these are actually antibodies to 7-globulin. The evidence is becoming overwhelming on this point. However, if anyone has evidence against the antibody hypothesis, this meeting is a good place to present it. 1. Physical properties. The rheumatoid factors have all the physical properties of classic 19s antibodies. Actually, more work has been dnne on rheumatoid factors in some ways than has been carried out on oth3er 19s antibodies; but that which has been done on such antibodies as 19s isoagglutinins shows a complete correspondence to rheumatoid factors in such physical properties as sedimentation coefficients, mobility, dissociation with sulfhydryl compounds, and all other properties that have been tested. 2. Antigenic properties. There is a complete correspondence between isolated rheumatoid factors and classic 195 antibodies such as the isoagglutinins. 3. The fine specificity for 7-glabulin of different genetic types. Characteristic of many rheumatoid factors, this answers the specificity criterion of antibodies. 4. Specificity for diferetzt portions of the y-globulin molecule. If these factors have specificity for more than one site on the molecule, it is much more difficult to conceive of them not representing antibodies produced against 7-globulin. The chances are very remote for a foreign antigen to cross-react with two widely divergent sites on y-globulin. The evidence is accumulating that there is multiple specificity at least as concerns the fragments produced by papain. The slow migrating fragments are easily separated from the fast fragments of 7-globulin and show complete antigenic differences. There are factors in rheumatoid arthritis sera that react with both fragments. 5. Heterogeneity. The multiplicity of factors favors the antibody hypothesis. Molecular heterogeneity (19s and 7s r-globulin) and the species heterogeneity are characteristic of classic antibodies. 6 . Production in experirnmtal animals. This does not as yet add too much to the concept but it has not ofFered any evidence to the contrary. Considerable research is now in process and conclusions soon will be appearing. Although evidence is overwhelming that the rheumatoid factors are antibodies, the question of whether they are autoantibodies is more difficult to 414 415 STRUCIVRE OF RHEUMATOID FACTORS U I 3 0 I ~ 5 l 7 9 l II 13 I 15 l 17 Fraction number P r o t e i n curve C o o t : whole a n t i - C D Ripley C o o t : pepsin digested anti-CD Ripley Fig. 1.-Density gradient separation of a rheumatoid arthritis serum. answer. One point to consider is that of isospecificity which Drs. Fudenberg, Harboe and others have shown for certain of the rheumatoid factors. How is it possible for these factors to show genetic specificity for y-globulin that is not present in that individual but is present in other individuals? Dr. Burnet has been very interested in this isospecificity and believes that this evidence fits with his ideas on clonal selection where somatic mutation in the cell lines cause increased proliferation of a certain cell-liberating antibody not of the standard pattern with active groups corresponding to patterns not found in that individual. This might be predicated on his theory of multiple specificities being innate in the individual before antigenic stimuli, with a somatic mutation giving rise to proliferation that then produces this foreign type of anti-y-globulin. However, one should note the autospecificity that does exist in the vast majority of these rheumatoid arthritis sera as has been demonstrated in the Gm system by Drs. Harboe, Fudenberg, and others. In many instances the autospecificity can be demonstrated in the complex formation with the patient's own y-globulin which is so characteristic of the ultracentrifuge pattern of these sera. These observations also indicate that one does not need denatured yglobulin or aggregated 7-globulin; there is direct interaction with the patient's own y-globulin and other 7s y-globulins. Figure 1, from work primarily by Drs. Osterland and Harboe, indicates a 416 IMMUNOLOGIC ASPECTS OF RHEUMATOID ARTHRITIS AND SLE Fig. 2.--Sera having large amounts of intermediate complexes, pointing out some of the more significant chemical features. simple density gradient separation of a rheumatoid arthritis serum. When one tests these fractions with whole Rh antibody on red cells, all the agglutinating activity is found in the bottom fractions corresponding to the 19s portion. However, when one splits the Rh antibody with an enzyme such as pepsin and coats the cell with the fragment of the Rh antibody, a whole new spectrum of anti-7-globulin factors becomes apparent. Here the activity with the pepsin-split Rh coat corresponds to the 7s fraction. A very good titer was obtained with this 7s rheumatoid factor which does not show itself at all with the whole Rh antibody on the red cell. There appears to be some buried determinant that is revealed by means of the enzymatic splitting. This has been the clearest demonstration of a 7s factor reacting with y-globulin in rheumatoid arthritis sera that we have had. This raises again the nomenclature question. Should all these agglutinators be called rheumatoid factors? DR. TOMASI:We have been interested in the question of whether a 7s type of rheumatoid factor exists. Dr. Kunkel pointed out sometime ago that certain sera from patients with rheumatoid arthritis contained a significant amount of complexes, designated intermediate complexes, which have sedimentation rates between 7 and 19S, usually in the 10 to 15s range. He suggested the possibility that these might represent the interaction of the 7s rheumatoid factor with other 7s 7-globulin molecules. Dr. Chodirker and f reexamined the question of the rheumatoid factor activity, using mainly the latex reaction, of these intermediate complexes. In figure 2 you will note sera having large amounts of intermediate complexes, pointing out some of the more significant chemical features. Frame A shows the native serum containing the intermediate complexes; and in the wedge window at the top is the same serum dialyzed to pH 3.5. The complexes have been dissociated by the acid. STRUCTUHE OF R5EUMATOID FACTORS 417 Fig. 3.-Latex fixation tubes positive. Dissociation begins around pH 4.5 which is approximately the pK of a carboxyl group which may be involved in the association reaction. On the alkaline side dissociation occurs around pH 10.5. It may be, however, that specific electrostatic interactions are not involved and that the subunits are bound together by closely fitting hydrophobic surfaces. Dissociation occurs at these pH’s because of the formation of highly charged units which result in dissociation when the electrostatic repulsions become greater than the attraction forces. Frame B shows the return of the complexes when the sera have been dialyzed to pH 3 and then back to pH 7. Frame C shows the complexes which have been treated with tenth-molar beta-mercaptoethanol.They are not dissociated, indicating that the covalent disulfide linkages are probably not involved here. We also examined these sera by three technics chosen for their potential in demonstrating a size distribution of activity: the density gradient technic, ion-exchange chromatography on DEAE cellulose, and the effect of betamercaptoethanol on rheumatoid factor activity. Figure 3 summarizes our studies with eight sera which can be divided into two general groups. The first three sera contain large amounts of intermediate complexes. The next three are sera with similar latex titers which have no intermediate complexes. The last two are sera from cases of Laennec’s cirrhosis and Waldenstrom’s macroglobulinemia,both showing high latex reactivity. The first group, represented by the first three sera with intermediate complexes, showed rheumatoid factor activity in the intermediate and 7s por- 418 IMMUNOLOGIC ASPECTS OF RHEUMATOID ARTHRITIS AND SLE tions of tlie density gradient and also in the pre-albumin and 7s regions of the chromatogram. The other sera showed latex activity only in the 19s region. The last column, “sulfhydryl reduction,” shows that the sera with the intermediate complexes lose approximately half of their latex reactivity in tenth-molar beta-mercaptoethanol. With the other sera the latex activity is completely abolished. These complexes, therefore, behave much like the 22s complexes resulting from reaction of 19s rheumatoid factor and 7s y-globulin. We believe that the intermediate complexes probably represent the interaction of 7s rheumatoid factor with 7-globulin and that the situation is analogous to that found with certain well defined macroglobulin antibodies which have a 7s counterpart. DR. EPSTEIN: Can Dr. Kunkel’s observation of a hidden antigen detecting a 7s material be equated with what you have just presented? You indicate that the patient’s own 7s 7-globulin has this exposed site because it can react with a 75 rheumatoid factor anti-y-globulin.Also, the material reacting with the pepsin treated anti-D material in Dr. Kunkel’s studies should then be positive in the latex test. DR. KUNKEL: I do not think a specific comparison has been made, but I think that the factors with which Dr. Tomasi is dealing are totally different from those we described. DR. EPSTEIN:Then there would be two types of 7s material: one capable of reacting with “native” 7s and another capable of reacting not with the patient’s native 7s but only after a site has been exposed? DR.KUNKEL:That seems most likely. DR. FRANKLIN: Shortly after Dr. Kunkel suggested the biologic activity of intermediate complexes, we encountered a patient whose complexes associated with a cryoglobulin which could be purified by exposure to cold and then isolated. The patient appeared to have a collagen disease; however, even after a postmortem examination the exact disease was not determined. Figure 4 shows the ultracentrifuge pattern of a 1:4 dilution of the patient’s serum on top and the same serum with a 19s macroglobulin peak as a marker on the bottom. As can be seen, there is a large series of complexes of very rapidly sedimenting molecules with S rates ranging from 10 to 45. Figure 5 shows results of some of the studies performed on the patient’s isolated material. The isolated cryoglobulin at pH 7 has a 7s peak and again a hetergeneous or rapidly sedimenting peak. As Dr. Tomasi just noted, this was completely dissociated at pH 4.3. On the other hand, mercaptoethanol reduced the size of the polymers at little to 10 to 15s peaks but did not completely reduce them to the 7s units. Finally, the lowest pattern on the right is the precipitate made with aggregated 7-globulin and redissolved at pH 4.3. As you can see, there is a very homogeneous 7s peak in this case and no 19s peak. The aggregates that were present in this precipitate have already sedimented to the bottom. It would appear that the isolated rheumatoid factor and the isolated rheumatoid factor precipitate from this patient were indeed 7s 7-globulins, as confirmed also by their antigenic properties and hexose content. STHUC'lWHE OF REEUMATOID FACTORS 419 Fig. 4.-Ultracentrifuge pattern of serum containing large amounts of rapidly sedimenting 10-40S complexes (top) and the same serum with a 19s macroglobulin added as a marker (bottom). Sedimentation proceeds from left to right. Together with Dr. Angelo Taranta, we also examined this serum for the ability to fix complement when it reacts with aggregated y-globulin. Neither the control sera nor the high titer 19s rheumatoid factor sera fixed complement with aggregated y-globulin. But when this patient's serum was added to aggregated y-globulin, particularly at low 7-globulin concentrations, there was fixation of about 20 units of hemolytic complement out of 80 that were added. This is the only time we encountered this observation in the large number of 18s rheumatoid factor sera that we have studied. Table 1 summarizes the findings of studies of this patient's serum compared to the usual 18s rheumatoid factors. It will be noted that the rheumatoid factor exists as a series of 10 to 40s complexes, which, unlike the 22s complexes usually seen, are not sensitive to mercaptoethanol. On the other hand, at pH 4.3 it is degraded to 7s units and loses its biologic activity; whereas classic rheumatoid factor is relatively stable at pH 4 and still exists as a 19s and 7s peak. These findings were confirmed by density gradient centrifugation which had to be done at pH 4.2 because it was a cryoglobulin. When the fractions thus obtained were dialyzed back to pH 7, the upper fractions, namely the 7s peak, retained biologic activity while the 19s peak was inactive. After dialysis at pH 7, intermediate complexes were again seen in the active fraction. Finally, immunologically and chemically this protein resembled 7s yglobulin rather than 19s 7-globulin. 420 IMMUNOLOGIC ASPECTS OF RHEUMATOID ARTHRITIS AND SLE Fig. 5.-Ultracentrifuge patterns of: ( 1 ) purified cryoglobulin in 0.9 per cent NaCl; ( 2 ) purified cryoglobulin in pH 4.3 glycine buffer; ( 3 ) purified cryoglobulin in 0.1 M mercaptoethanol; ( 4 ) precipitate made with aggregated y-globulin dissolved in pH 4.3 glycine buffer. The aggregates have sedimented to the bottom and only a homogeneous 7s peak is visible. DR.RAGAN:Dr. Kunkel, how frequently did reactivity of the 7s component occur among rheumatoid sera? DR. KUNKEL: The distribution of the buried determinant factor is somewhat different from the classic rheumatoid factors. We found that approximately 50 per cent of rheumatoid arthritis patients showed this factor and some of them in extremely high titer. DR. RAGAN:Was it found in patients who did not have high titer of ordinary rheumatoid factor? DR. KUNKEL:Yes, and there was no correlation between the two. It was also found in approximately 20 per cent of normal sera, usually at low titer. 421 STRUCTURE OF RHEUMATOID FACTORS Table 1.-Properties of 7s Rheumatoid Factors Reacting with Human 7s Gamma Globulin Compared to 19s Rheumatoid Factors 19s Rheumatoid Factor 7S Rheumatoid Factor Procedure Native RH 3-4 SH '7s y added Sed.Rate RF Activity Other Sed. Rate 22s 19s+7s 75 22s 19s y added Den. grad. pH 4.2 Immunologic Hexose Comolement fixation 7s Fx 19SFx RF Activity ++++ ++++ ++++ Other (-) no change (--) +++ 19s y 5.5% No Dr. Williams also found it in many of his bacterial endocarditis sera where it appeared during the acute endocarditis and disappeared following treatment. DR. EPSTEIN:The procedure here tested against rheumatoid serum would also detect anti-Inv determinants because that would be included in the pepsin fragment coated cell. Was it checked to see that this was not a 19s anti-Inv rather than a 7 s material? h.HARBOE:It was settled in two ways. The anti-Inv's we investigated were all 19s. The new factors were, except in two cases, only 7s. Inhibition experiments show that they did not have anti-Inv specifkity. However, the anti-Inv system worked with the pepsin Rh coat. DR. HErMER: In the past, we had thought all rheumatoid factors to be euglobulins. Recently Dr. Nosenzo and I obtained serum from a patient with macroglobulinemia of which 50 per cent was 19s macroglobulin and could be absorbed out with denatured y-globulin. On euglobulin fractionation we were much surprised to find 50 per cent of the rheumatoid factor in the pseudoglobulin fraction. Dialyzing this fraction against .001 molar Tris buffer at pH 7 or phosphate buffer around pH 5.5, this rheumatoid factor when chromatographed on DEAE cellulose was 19s. It came off the DEAE column practically free of any other contaminant. This material gave a positive latex test but was completely negative when tested by sheep cells sensitized with rabbit amboceptor. DR. KUNKEL:This appears similar to a case presented by Dr. LoSpalluto in which he got direct interaction between 19s and 7 s 7-globulins in a cryoglobulin precipitation reaction without using aggregates.l Did you get cryoeuglobulin properties when you added 7s y -globulin? We did not do that. DR. HEXMER: DR. LOSPAUUTO:In the case just referred to by Dr. Kunkel, the 19s yglobulin was a euglobulin which precipitated rather readily. We could isolate it in fairly pure form, but once isolated it did not have the property of precipitation in cold. It behaved like a rheumatoid factor in that one could precipitate it with denatured 7-globulin. This was not the reversible type of cryoglobulin; it did not dissolve on warming. The formation of the complex with 7s y-globulin resulted not in 422 IMMUNOLOGIC ASPECTS OF RHEUMATOID ARTHRITIS AND SLE precipitation at room temperature but only in the cold. Thus, the solubility characteristics appear to be different. DR. KUNKEL: Not necessarily. In the whole serum you had 7s 7-globulin initially, and so it would be a cryoeuglobulin as long as it is a complex; as soon as you free it of the 7s 7-globulin it is no longer a euglobulin. I stress t h i s point because we have had several sera that behave this way which we thought were analogous to the case of Dr. LoSpalluto. One of thsese, which has been studied extensively by Dr. Pernis in Italy, gives a precipitin curve with 7 s 7-globulin when the tubes are put in the cold but not at room temperature. It shows solubility in excess 7-globulin. Others of our cases also show this phenomenon but only after lowering the pH. They precipitate with added 7-globulin in the cold at the lower pH. This occurs more readily under the conditions giving rise to euglobulin precipitation. This is apparently a rather widespread phenomenon, that one has this precipitin reaction directly with 7s 7-globulin when you have the isolated factor free of 7 s 7-globulin. However, we had certain difficulties following this particular cryoglobulin. We wanted to see if the addition of excess 7 s to the isolated cryoglobulin approximated 50 per cent each of 7 s and 19s. We tried to reconstitute a complex by increasing the concentration of 7 s 7:globulin but were unsuccessful. We did get a cryoglobulin but not the soluble complex as it looked originally. I suspect that there might be some specificity as regards the 7s fraction. DR. FRANKLIN: I would like to point out the frequency of Dr. LoSpalluto’s observations. Since he reported his original paper, we have seen five such cryoglobulins which were selected out of a total of 15 or 16 that we examined. Four of these sera were selected at random from 15 or 16 cryoglobulin, that Dr. Meltzer and I have studied and the remaining one was sent us by Dr. Waldenstrom. The interesting finding is that none existed as a 22s complex in the serum. On the contrary, all of them existed in the whole sera and after isolation as separate 19s and 7s peaks, both of which were required to precipitate in the cold. The 19s fraction alone was able to precipitate with aggregated 7-globulin. DR.ZIFF: What would happen if you tried to reconstitute using the original patient’s own 7S? DR.FRANKLIN: It works when you take the patient’s own or homologous 7 s 7-globulin. Also, if you take papain fragment B of 7-globulin it will again become a cryoglobulin. We could never get a 22s complex in these patients. They have always existed as 7s and 19s peaks in the serum and isolated cryoglobulin. DR.BLOCH:These observations may explain the results of experiments in which rheumatoid factor in sera from patients with systemic lupus erythematosus is distinguished from that occurring in rheumatoid patients by differences in solubility. Svartz and Schlossman,2 as well as Black et al.,3 have shown that rheumatoid factor in most SLE sera is not precipitated by the addition of 14 volumes of cold distilled water. Rheumatoid factor occurring STRUCTURE OF RHEUMATOID FACTORS 423 in sera from rheumatoid patients is precipitated under these conditions. SLE sera may contain insufficient amounts of the appropriate 7s 7-globulin for complex formation. DR. KUNKEL:One important point regarding precipitin reaction is that the solubility of rheumatoid factor as it circulates may be somewhat controlled by the amount of 7s y-globulin present. Perhaps this is important in arteritis. If there is a lower concentration of 7-globulin for various reasons, you will get a decreased solubility and precipitation. DR. ZIFF: Then people with low gammas ought to get arkritis? DR. KUNKEL:I think that is a possibility. DR. CHRISTIAN: Do you imply that the only 19s which comes down in a euglobulin is complexed with 7S? DR. KUNKEL:I think in some cases it is true that it is a complex that makes the euglobulin. DR.CHRISTIAN: This would not be true of isohemagglutinins and the various bacterial antibodies that separate as euglobulins. DR. KUNKEL: Well, many of these are not typical euglobulins. The rheumatoid factor as it is measured is much more insoluble than, for example, the 19s isoagglutinins. DR. EPSTEIN:Do you visualize the 22s material in the euglobulin to be an antigen-antibody complex which partially dissociates when the euglobulin is dissolved in saline? I ask this because, using gel filtration to obtain the euglobulins, there appears to be a direct correlation between the titer of the whole serum and the amount of 22s material in the e~globulin.~ Such complexes are found in the euglobulin even when none were observed in ultracentrifuge study of the whole serum. It would be my feeling that at least a potential to form 22s complexes exists in all rheumatoid sera. DR. KUNKEL: I wonder if anyone would like to discuss the question of evidence against the hypothesis that the rheumatoid factor is antibody. DR. HEIMER:Our laboratory might have suggestive evidence that the rheumatoid factors are not necessarily antibody. There appear to be differences in the behavior of rheumatoid factor in complement fixation reactions. One can elicit two different types of response depending on whether one uses soluble antigen-antibody complexes or y-globulin coated on latex particles. Highly purified rheumatoid factor does not fix complement. Rheumatoid factor also fails to disturb the complement uptake by soluble antigen-antibody complexes. In other words, the antigen and antibody complex contains separate sites for complement fixation and the rheumatoid factor fixation and these sites do not interfere with one another. In this type of system the rheumatoid factor behaves like an antibody; it has its site of attachment separate from that of complement. In the other system, F-I1 coated latex particles which also fix complement, the addition of rheumatoid factor results in complete inhibition of complement fixation. In this system complement and rheumatoid factor either compete for the same site or for perhaps neighboring sites. Now, if they com- 424 lMMUNOLOGIC ASPECTS OF RHEUMATOID ARTHRrTIS AND SLE Pete for the same site, then the rheumatoid factor is not behaving like an antibody but like an inactive component of complement. DR.KUNKEL:Why shouldn't it be to the same site as complement? DR. HEIMER: I have no proof. I have not been able to displace one with the other. DR.MILGROM:In the F-I1 latex system, complement is not fixed by antigenantibody complexes, rather it is fixation of complement by aggregated 7globulin and that may be a different phenomenon. DR. HEIMER:Yes, but rheumatoid factor inhibits this reaction. Of course, one other objection to this would be that 7-globulin absorbed on the latex particle has a different conformation from the 7-globulin joined to antigen. DR. VAUGHAN: Dr. Heimer, that F-I1 is human y-globulin is it not? And the antibody in your antigen-antibody complexes is rabbit 7-globulin? DR. HEIMER: Yes, and this is certainly one further objection to the scheme. DR. KUNKEL: We could well get lost in the complement story. It is so mysterious why some 1% antibodies fix complement and some do not, we get into an impossible situation when seeking this as evidence for the antibody hypothesis for the rheumatoid factor. DR. EPSTEIN: One of the objections to the antibody hypoth'esis has been the duration and lack of variation of the level of antibody correlated with disease activity. Rheumatoid arthritics who manufacture this antibody rheumatoid factor may do so in essentially unchanging amounts over several decades. Assuming this is due to an antigenic stimulus, the antigen would have to be constantly present over this same period. It has been demonstrated that following a single antigenic stimulus, rabbits may produce high levels of hemagglutinating antibcdy for periods far beyond those associated with detectable precipitating antibody." DR. KUNKEL: The point you make is why is there so prolonged a 19s response when we know in experimental animals it is the initial stage of antibody formation that is 19s which then falls off? We do have the other example of the isoagglutinins where on one stimulus we have increased 19s isoagglutinins that go on through the whole life of the individual. Just what this means I do not know. DR. BENACERRAF: Have patients been investigated for long enough periods to make certain that their pattern of serologic reactivity has neither changed in the antibody types nor in the specificity which they show? Is it the same when you get a patient the first time or 2 or 3 years later? The fact that they do have some antibodies has no meaning in the relationship to the variety which these may take over a period of time. I wonder whether the disease process has been studied in this respect. DR. ZIFP: I think the titer tends to stay the same. DR. SINGER: We had about 100 sera over the past few years in which we observed no changes in titers with the latex with the exception of 2 patients. DR. BENACERRAF: What about immunologic specificity? The spectrum? DR.ZIFF: Most tests have been done with aggregated human 7-globulin. DR. MIESCHER:With the hypothesis that the rheumatoid factor is formed STRUCTURE OF RHEUMATOID FACTORS 425 Fig. 6.-Variations in amount of intermediate complexes over period of time in serum from rheumatoid arthritis patient. due to stimulation by altered autologous 7-globulin, altered in the sense of interaction with an antigenic substance, if the patient would be exposed throughout his whole life to antigenic substances, it might explain why he continues to form this type of rheumatoid factor based on a special genetic pattern. Following this thinking, is it possible that the so-called 22s complexes with autologous 7-globulin are with altered 7-globulin? Did you investigate whether the rheumatoid factor reacts better with an autologous antibody after it has reacted with an antigen? For example, did you make an antigen-antibody react with an autologous rheumatoid antibody, dissociate it again at acid pH and then see if this dissociated antibody will better react with the rheumatoid factor than before its interaction with an antigen? I think the extraordinary thing is that it is a 19s and Dn. MCDUFFIE: stays there so long when what we would expect to find is 7 s . Even in the experiments with BSA and other antigens, where it has been shown that their early antibody is 19s and the later antibody is 7S, we do not know if the specificity of the two is the same. In studying 19s and 7s anti-blood type antibodies several years ago, we noticed quite a marked difFerence in the inhibitory power of human versus animal blood group substances. Furthermore, with the sheep cells the difference between the heterophil and isophil antibody with time might also suggest that certain antigenic groups stimulate 19s and not 7s antibody production. DR. TOMASI: We have a little data on the variation in the amounts of the intermediate complexes with time. Figure 6 illustrates the important point that the amount of these complexes can vary significantly over the course of time. This serum is from a patient with intermediate complexes and rheumatoid arthritis. There are four samples of serum. The time interval between the second and third sample, in which the intermediate complexes have almost disappeared, was about 6 months. The return of complexes occurred over the next 6 months. We observed another patient in which the changes in the amount of the 426 IMMUNOLOGIC ASPECTS OF RHEUMATOID ARTHRITIS AND SLE intermediate complexes, without changes in therapy, were just as significant as this. D R . HOLMAN: Were there any changes in disease activity? D R . TOMASI: There were no changes in disease activity but there was considerable change in total latex titer. The first sample had a latex titer of 1 million and the third sample was about 1:20,000 in the first patient mentioned. DR.RAGAN: I think the question of change in titer related to disease activity must be answered with difficulty. Most indications are that the titer diminishes as the disease gets better when measured using human 7-globulin as the antigen. Those that have measured using sensitized sheep cells say almost unanimously they find no change in titer. We know that in liver disease, as Dr. Kunkel and others have pointed out, and in other diseases, you get the so-called latex type of rheumatoid factor (reaction restricted to human y-globulin), However, I think it might help clear our thinking if we restrict the use of the term rheumatoid factor to that which reacts with sheep cells or rabbit 7-globulin and use some other term for that which reacts with human 7-globulin. The question of nomenclature should be discussed as with all DR.KUNKEL: these different anti-7-globulin factors there is a real question as to whether they should be called anti-7-globulin factors rather than rheumatoid factor. I think it would be better to use the term anti-7-globulinfactor more universally. REFERENCES 1. LoSpalluto, J., Donvard, B., Miller, W., Jr., and Ziff, M.: Cryoglobulinemia based on interaction between a gamma macroglobulin and 7s gamma globulin. Am. J. Med. 32:142, 1962. 0. Svartz, N., and Schlossman, K.: Agglutination of sensitized sheep erythrocytes in disseminated lupus erythematosus. Ann. Rheumat. Dis. 16:73, 1957. 3. Black, A., Goldin, M., Poske, R. M., and Malmed, L.: Differentiation between rheumatoid arthritis and systemic lupus erythematosus by sheep cell agglutination tests. Arth. & Rheumat. 2: 99, 1959. 4. Epstein, W., and Tan, M.: Gamma globulin interactions in the sera of 2 patients with rheumatoid arthritis studied by gel filtration. J. Lab. & Clin. Med. 60: 125, 1962. 5. Blumer, H., Richter, F., Cua-Lim, and Rose, B.: Precipitating and nonprecipitating antibodies in the primary and secondary immune responses. Rate of decline, anaphylaxis-sensitizing capacity and the effect of cortisone. J. Iminunol. 88:669, 1962.