Instantaneous roll-blot from cellulose acetate after electrophoresis. A versatile tool for monoclonal antibody characterization.

An instantaneous blotting method from cellulose acetate to nitrocellulose was developed using a high pressure roll apparatus. This method was applied to the early characterization of monoclonal antibody specificity and to monoclonal immunoglobulin typing in mouse hybridoma supernatants, human sera or unconcentrated urines. Immunoglobulins were then revealed using, successively, anti-isotype specific monoclonal or polyclonal antibodies, avidin-biotinylated peroxidase complexes and cobalt-enhanced diaminobenzidine substrate.

[Hemorrhagic fever with renal syndrome due to Hantaan virus or a serologically related virus]. / Fièvre hémorragique avec syndrome rénal due au virus Hantaan ou à un virus sérologiquement apparenté.

Seven cases of acute renal failure consecutive to haemorrhagic fever with renal syndrome (HFRS) due to the Hantaan virus or to a serologically related virus are reported. These cases were observed in north-eastern France between March, 1983 and January, 1984. All patients were of rural origin and had been in contact with field mice. The predominant initial clinical symptoms were signs of infection and diffuse muscle pain, without evidence of haemorrhage. However, massive proteinuria was noted, and acute anuric renal failure unaccompanied by oedema or arterial hypertension developed. Renal biopsy performed in 2 patients showed tubular and interstitial nephritis but no glomerular or vascular lesions. Two patients only required haemodialysis. All patients recovered within 2 to 8 weeks without sequelae. Antibodies directed against the Hantaan virus were detected by indirect immunofluorescence tests, and seroconversion could be demonstrated in 2 patients seen at a sufficiently early stage. The risk of epidemics suggested by this small outbreak of HFRS can only be evaluated after an exhaustive epidemiological study.

[Strict correlation between anti-RANA antibodies and anti-EBNA antibodies apart from rheumatoid arthritis. Assessment of the diagnostic value of anti-RANA antibody]. / Corrélation étroite entre l'anticorps anti-RANA et l'anticorps anti-EBNA en dehors de la polyarthrite rhumatoïde. Critique de la valeur diagnostique de l'anticorps anti-RANA.

In order to study the relationship between anti-RANA antibodies (aRANA) and the antibodies capable of recognising other Epstein-Barr induced antigens, we examined the sera of 51 patients without rheumatoid arthritis for anti-RANA, anti-EA, anti-VCA, and anti-EBNA antibodies. These subjects were cases of Burkitt's lymphoma, infectious mononucleosis, naso-pharyngeal carcinoma, immune disorders, Hodgkin's disease and normal controls. A blind study was conducted in two separate laboratories. Anti-RANA was detected by indirect immunofluorescence on RAJI cells synchronised in phase G1. There was a very strict correlation between anti-RANA and anti-EBNA with no correlation between anti-RANA and anti-EA or anti-VCA. In another experiment, 15 coded sera from patients with classical rheumatoid arthritis were tested following adsorption of the rheumatoid factor. One serum was found to be devoid of both anti-RANA and anti-EBNA. These results demonstrate that anti-RANA is widely distributed outside of rheumatoid arthritis and they question the distinction between EBNA and RANA, as the antibody titres directed against these antigens are regularly linked together.

Detection and measurement of rheumatoid factor using a new immunoenzyme technique with peroxidase/anti-peroxidase complex.

A new, solid-phase enzymatic technique is proposed for the detection and measurement of rheumatoid factors (RF). A complex (PAP) consisting of peroxidase and rabbit IgG anti-peroxidase antibodies was used as antigen. The sensitivity of the technique is such that even "physiological" levels of RF may be detected in the majority of serum samples. The intra-assay and inter-assay repeatability (6-8% and 12%) is better than that for either of the traditional agglutination techniques (latex fixation test and Waaler-Rose reaction, or LWR) and this means that the serological evolution of the illness may be followed more accurately for each patient. The detection of non-agglutinating RF was also possible. The antigen used (i. e. rabbit IgG) was fixed immunologically to the enzyme, hence it was not denatured by chemically labelling. This antigen resembles that of the Waaler-Rose reaction. A variation of this method enabled the detection of RF immunoglobulin class, when only the IgM fraction of the sera tested could be fixed to the solid phase by means of anti-mu F(ab')2 antibody before introduction of the antigen. The affinity constant could be measured without antibody purification. In 211 cases of adult rheumatoid arthritis, abnormally high levels of RF were revealed in 78% of the subjects using the PAP technique and in only 53% with the agglutination (LWR) techniques. On the other hand, the RF levels were within normal limits in the 51 cases of other inflammatory rheumatisms lacking RF according to LWR techniques, in 62 cases of non-inflammatory diseases (hip osteoarthrosis, low back pain) and also in the 42 cases of other inflammatory diseases.

Immunological study of hydatidosis. I. Evaluation of immunoelectrodiffusion tests and enzyme-linked immunoelectrodiffusion assay (ELIEDA) in human hydatidosis.

The comparative sensitivity, specificity and rapidity of immunoelectrodiffusion (IED) on cellulose acetate membranes and of immunoelectrophoresis (IEP) on agarose gel, were evaluated in the diagnosis of hydatidosis. Pooled crude fluid from several cattle hydatis cysts was used as antigen in tests on 1,750 non-hydatis and 400 hydatis sera obtained from patients with hepatic, pulmonary, splenic, peritoneal and cerebral hydatidosis before and after surgery. Coupled to a specific human reference serum for arc 5, IED shows a specificity comparable to that of IEP but its sensitivity is slightly higher and the amount of antigen needed is very small. The appearance of a typical "gloved finger" pattern in sera from patients with ruptured cysts emphasizes the interest of quick results (3 hours) obtained by this method. In order to increase the sensitivity of IED and to define the class of immunoglobulins involved in the antigen-antibody reaction, we have coupled this method to an enzymatic technique. The immune complexes precipitated by IED were treated with peroxidase-labelled antibodies specific to each class of human immunoglobulins. The specificity of this enzyme-linked immunoelectrodiffusion assay (ELIEDA) permits one to follow the immunologic evolution of hydatidosis and to identify IgM in ruptured hepatic cysts and IgA in pulmonary cysts.

[Renal lesions in congenital syphilis. A case report]. / L'atteinte rénale de la syphilis congénitale. A propos d'une observation.

The authors describe a case of congenital syphilitic nephropathy in an infant. The review of the reports in the literature shows many similarities of this congenital disease with the syphilitic nephrotic syndrome in the adult. The prognosis is favorable provided the diagnosis is made sufficiently early. Histological studies utilizing light-, electron- and immunofluorescence-microscopy gave results consistent with extramembranous glomerulonephritis with possible evolution towards endo- and extracapillary proliferative glomerulonephritis. This is one of the rare renal diseases with a known antigen.

[A critical study of immuno-enzymatic reactions coupled with the precipitation tests on cellulose acetate membranes]. / Etude critique des réactions immuno-enzymatiques couplées aux tests de précipitation sur membranes d'acétate de cellulose.

Precipitating tests carried out on cellulose acetate membrane can be increased by treating the immune complexes with enzyme linked anti-immunoglobulin antibodies. From our trials in parastic diseases (Amoebiasis, schistosomiasis, fasciolasis, filariasis, hydatidosis, trichinosis) and mycosis (Aspergillosis, Candidiasis), it seems that the immuno-enzymatic labelling of the "active" precipitating reactions should only be taken in consideration. We must insit on the importance of ELIEDA (enzyme-linked-immuno-electrodiffusion-assay) and ELIDEPA (enzyme-linked-immuno-double-electro-phoresis-assay). Both of these analytical assays are particularly sensitive. The sequence of appearance of the multiple precipitating systems of the complex parasitic mosaic can easily be watched. The different classes of immunoglobulins and their kinetics are determined by the use of monospecific antibodies linked to different enzymes which give a polychromic specific staining.

[Precipitating immunoenzyme revealed tests on cellulose acetate. Use of ELIEDA (enzyme-linked-immuno-electro-diffusion assay) and ELIDEPA (enzyme-linked-immuno-double-electrophoresis-assay) in parasitology]. / Etude critique de la révélation immuno-enzymatique des réactions d'immuno-précipitation sur acétate de cellulose. Intérêt d'ELIEDA (enzyme-linked-immuno-electro-diffusion-assay) et d'ELIDEPA (enzyme-linked-immuno-double-electro-phoresis-assay) en parasitologie.

Precipitating tests carried out on cellulose acetate membrane were revealed by treating the immune complexes with enzyme linked anti-immunoglobulin antibodies. We insist on the importance of ELIEDA (enzyme-linked-immuno-electro-diffusion-assay) and ELIDEPA (enzyme-linked-immuno-double-electro-phoresis-assay) for the study of antibody classes involved in the immune response.

The use of ELIEDA (enzyme-linked-immuno-electro-diffusion-assay)) in the study of humoral antibodies in human schistosomiasis.

In order to improve the sensitivity of immuno-electrodiffusion (IED) we have developed a technique of treating the precipitated immune complexes with enzyme-labelled antibodies. This technique we term ELIEDA (Enzyme-Linked-Immuno-Electro-Diffusion-Assay). The enzyme-labelled antibodies can be made monospecific for each class of immunoglobulins found in patients with schistosomiasis and which precipitate with schistosome antigens. We believe that this sensitive technique will prove valuable in studying the sequential development of antibodies in human and experimental schistosomiasis.

[Exploration of humoral immunity in aspergillosis: value of enzyme linked immuno-electro-diffusion assay]. / Exploration de l'immunite humorale dans l'aspergillose: intérêt de l'E.I.I.E.D.A.

In order to increase the sensitivity of the method of immunoelectrodiffusion (I.E.D.) and to isolate the class of immunoglobulins implicated or involved in the Aspergillose antigen-antibody reaction, we supplemented the I.E.D. by combining or incorporating it with an enzymatic technique. By this method the immune complexes precipitated are taken up by antibodies conjugated to one enzyme and specific to the immunoglobulins of each class. This constitutes the technique of E.L.I.E.D.A. (enzyme-linked-immuno-electro-diffusion assay). The sensitivity and the specificity of the proposed method would allow us to follow the qualitative evolution of the aspergillar antibodies and would lend support to further arguments on the evolutionary character of the inflammatory response.

ELIEDA (enzyme-linked-immuno-electro-diffusion-assay): application of a combined immunoelectrodiffusion and immunoenzyme method to the study of immune response in parasitic infections.

The sensitivity of the technique of immunoelectrodiffusion (IED) has been enhanced by combining it with an enzyme method; this also enabled the class of immunoglobulins involved in the reactions to be determined by treating the precipitated immune complexes with enzyme-labelled class-specific antisera. The method has been employed in studies of the immunology of parasitic infections and these confirmed its sensitivity and specificity. ELIEDA provides a valuable tool for analysing responses to complex immunogens.

[Humoral study of human parasitic diseases. Interest of ELIEDA. (Enzyme--linked--immuno--electro--diffusion--assay) (author's transl)]. / Etude immunologique des parasitoses humaines. Intérêt de l'ELIEDA (Enzyme-linked-immuno-electro-diffusion-assay)

In order to sensitize the reaction of immuno-electro-diffusion (IED) and to define the class of immuno-globulins implied in the antigen-antibody reaction, we have completed the technique of IED by coupling it to an enzymatic technique. In such a way the immune complexes are treated by enzyme labeled antibodies specific to each class of immunoglobulin. This association realizes the technique of ELIEDA (enzyme-linked-immuno-electro-diffusion-assay). The sensitivity and the specificity of this method allow the follow up of the qualitative evolution of antibodies during natural or experimental parasitic diseases.

Immunological study of hydatidosis. I. Evaluation of the test of immuno-electrodiffusion in the humoral study of human hydatidosis.

The importance of immunological arguments in the discussion of hydatidosis, the permanent increase of requirements, the risk of cross reactions, the frequency of sero-epidemiological surveys imply the utilisation of specific and rapid micro-methods. The technique of immunoelectrodiffusion on a cellulose acetate membrane answers partly to those criteria; we have used it since 1971 in the diagnosis of parasitic infections and mainly to that of hydatidosis. Two hundred cases (400 samples) of the latter were studied longitudinally and reported. This micro-method is very specific (co-immunoelectrodiffusion), equal to that of immunoelectrophoresis. Its rapidity (3 to 4 hours) is close to that of indirect immunofluorescence. The revelation of suggestive diagrams of fissuration underlines the relative emergency of the definitive interpretation of an immunological examination in the diagnosis of hydatidosis.