Felix Milgrom

Human Sera with “Anti‐Antibody”

Out of 2000 human sera tested, 10 were shown to agglutinate O Rh + red cells sensitized with incomplete anti‐D antibodies. The reaction was apparently analogous to the reaction obtained with Coombs serum.

Evolution of membranous nephropathy into anti-glomerular-basement-membrane glomerulonephritis.

Abstract In a patient with biopsy-proved membranous nephropathy (nephrotic syndrome) acute fatal renal failure suddenly developed. Autopsy revealed rapidly progressive glomerulonephritis superimposed on the previous membranous changes. IgG eluted from the kidney was shown by immunofluorescence technic to bind to the glomerular basement membrane of normal human and monkey kidneys, and was capable of producing an acute anti-glomerular-basement-membrane nephritis in two monkeys. The IgG antibody, by radioisotopic methods, was specifically directed against primate kidney. It is postulated that immune complexes responsible for the original membranous nephropathy induced release of antigenic glomerular-basement-membrane fragments into the circulation, stimulating formation of antibodies to the membrane and producing fatal acute glomerulonephritis. (N Engl J Med 290:1340–1344, 1974)

Anti-basement membrane antibodies and antigen--antibody complexes in rabbits injected with mercuric chloride.

Abstract New Zealand rabbits were injected three times a week with 2 mg of mercuric chloride (HgCl2) (MC) to test the hypothesis that toxic agents release autologous antigens, generating antibody response and tissue injury. After 2 weeks of injections the rabbits developed anti-basement membrane antibodies binding to renal and extrarenal basement membranes and to the peri- and endo-mysium of skeletal muscles. Glomerular histology appeared normal. Membranous glomerulonephritis developed 1–2 months later with granular subepithelial deposits containing rabbit IgG and C3. Similar deposits were present in the basement membranes of other organs. Raji cell tests for immune complexes in the sera were negative in the initial stages and positive in the late stages of the disease. Radioactive MC was found in the cytoplasm of renal tubular, intestinal, and hepatic epithelial cells but not in glomerular deposits. Renal eluates and selected sera reacted with the basement membranes and the collagen matrix in normal or collagenase-digested sections of renal or extrarenal normal rabbit tissues. This reactivity was reduced or abolished by washing tissue sections in PBS or by absorption of renal eluates with whole kidney homogenates. The findings are consistent with the hypothesis that MC induces a biphasic disease characterized first by the production of antibodies to basement membranes and the extracellular collagen matrix and subsequently by antigen-antibody complexes formed in situ and in the circulation and presumably containing soluble polysaccharide components of the collagen matrix.

Heterophile antigens and antibodies in medicine.

When an immune serum reacts with cells, tissues, or body fluids of species different from that which supplied the material for immunization, it is generally assumed that these cross-reactions are due to the sharing of antigen(s) or antigenic determinant(s) among various species, as pointed out by Ehrlich and Morgenroth as early as 1901. The cross-reacting antibodies can be divided into two categories (1) those reacting only with antigens of closely related species — cross-reacting, species-specific antibodies and (2) those acting on antigens of unrelated species. Probably the first report to describe cross-reacting, species-specific antibodies was that of Ehrlich and Morgenroth in 1900. They observed that when rabbits were immunized with goat erythrocytes, the resulting antisera lysed sheep erythrocytes and vice versa. Forssman antibodies are the best known example of antibodies that act on antigens of unrelated species. In 1907, Frouin reported that rabbits immunized with chicken egg yolk formed antibodies that lysed canine erythrocytes. Furthermore, he and Lisbonne (1911) clearly showed that the rabbit anti-egg yolk sera could lyse erythrocytes of sheep, horse, and goat, all of which are now known as Forssman-positive species. In 1911, Forssman reported a similar observation that rabbits immunized with aqueous suspensions of guinea pig, horse, or cat tissues formed antibodies which lysed erythrocytes of sheep, a species apparently unrelated to any of those from which immunizing antigens were derived. Forssman named these antibodies heterologous antibodies (heterologe Antikorper).

Dissociation of immune complexes in tissue sections by excess of antigen.

Immune complexes (IC) present in the glomeruli of rabbits with chronic serum sickness (CSS) and in patients with systemic lupus erythematosus (SLE), idiopathic membranous nephropathy (IMN), and acute poststreptococcal glomerulonephritis (PSGN) were analyzed by incubation with antigenic preparations. The efficacy of these preparations to dissolve IC was assayed by comparison of results of direct immunofluorescence tests performed with the kidney tissues before and after incubation with antigenic preparations. The FITC-conjugated antisera used in these tests were specific for IgG, C3, and-in the case of CSS-for the eliciting antigen, bovine serum albumin (BSA). During the acute proteinuric phase of CSS in rabbits, incubation of tissue sections with BSA alone led to complete dissolution of IC. In many rabbits with late phase proteinuria, however, tissues had to be incubated with both BSA and aggregated fraction II of rabbit serum. In all biopsy specimens from patients with IMN, and in some specimens from patients with PSGN and SLE, aggregated fraction II of human serum resulted in complete or incomplete dissolution of IC. On the other hand, incubation of tissues with excess DNA in SLE or with streptococcal antigens PSGN did not lead to dissolution of IC. These studies suggest significant participation of antibodies to aggregated immunoglobulins (i.e., rheumatoid factors or rheumatoid-like factors) in IC found in the above-mentioned diseases. Other antigen -antibody systems, however, may also contribute to the deposits in the glomerulonephritides studied.

Detection of circulating immune complexes by the inhibition of anti-antibody.

Abstract An inhibition test of agglutination of sensitized erythrocytes by human or rabbit anti-antibody was established for the detection of circulating immune complexes. Anti-antibody is a previously described IgM serum factor which combined specifically with homologous IgG antibodies that underwent molecular transformation in reaction with their corresponding antigen. In testing in vitro -formed immune complexes by human or rabbit IgG antibodies, it was found that this test is capable of detecting immune complexes at antigen concentration as low as 2–3 μg/ml. After the addition of corresponding antigens, all 14 serum-sickness sera and all 9 chronic thyroiditis sera which contained precipitating antibodies inhibited human anti-antibody. IgG of normal human sera, heataggregated IgG, and repeatedly frozen and thawed normal sera gave negative inhibition tests. By means of this inhibition test, immune complexes were demonstrated in 35% of the systemic lupus erythematosus sera, 69% of the rheumatoid arthritis sera, 54% of the malaria sera, 22% of the postrenal transplantation sera, 22% of the liver disease sera, 32% of the leukemia sera, and 35% of the lymphoma sera. All 19 of the leprosy sera from India also gave positive results. None of the sera of 36 healthy staff members and only 3% of the sera of random blood donors gave positive results.

Rabbit Sera with “Anti-Antibody”*

A normal rabbit serum was incidentally found which agglutinated, up to a dilution of 1:128, rabbit blood group G erythrocytes sensitized by incomplete anti‐G sera of rabbit origin. A similar agglutinating activity was then demonstrated in the sera of 2.7% of apparently normal rabbits. The serological properties of the agglutinating factor were quite analogous to those of the factor previously described in human sera, called anti‐antibody. The rabbit anti‐antibody resisted heating up to 56°C and was associated with γ‐β‐globulin electrophoretic fractions of the serum. It could not be neutralized by any whole rabbit serum or by the γ‐globulin fraction of pooled rabbit serum. However, it was readily absorbed by immune complexes composed of any antigen and the corresponding antibody of rabbit origin. On the basis of experiments presented, the rabbit anti‐antibody could be considered as a specific agent capable of combining with rabbit immune globulin modified in the course of serological reaction but not with the free normal or immune rabbit γ‐globulin. The possible antibody character of the serum factor under investigation has been discussed.

Circulating immune complexes after kidney transplantation.

Sera from patients who had received renal allografts were studied for the presence of circulating immune complexes by using platelet aggregation technique combined with density gradient centrifugation. A simple and highly reproducible modification of the platelet aggregation technique, employing the use of relatively small amounts of blood from pretested donors as the source for platelets, is described. Immune complexes were detected in post-transplantation sera from 3 out of 16 patients. The development of a persistent immune complex state as a consequence of grafting was concluded in one patient.

DETECTION OF HUMAN LYMPHOCYTE ISOANTIGENS BY RABBIT HOMOTRANSPLANTATION SERA

SUMMARY Rabbit transplantation sera were studied for their cytotoxic activity against human lymphocytes. Of 41 skin graft sera tested, 15 gave definite positive reactions with some lymphocyte samples. The family studies showed that the human lymphocyte isoantigens recognized by rabbit transplantation sera are inheritable, and they appear to be controlled by a single chromosomal locus which seems to be closely linked or identical with the HL-A locus

Immunologic sequelae of thermal injury.

1) Thermal injury induces a marked inhibition in the ability of tuberculin‐sensitive guinea pigs to respond to intradermal challenge with tuberculin. The effect is most marked during the first and second weeks after burning, but may persist as long as five weeks.