[Immunoenzymatic determination of free immunoglobulin light chains in the cerebrospinal fluid: its value in the biological diagnosis of multiple sclerosis]. / Dosage immunoenzymatique des chaînes légères libres d'immunoglobulines dans le liquide céphalorachidien: intérêt dans le diagnostic biologique de la sclérose en plaques.

Based on previous data on free immunoglobulin light chains in the cerebrospinal fluid (CSF) of patients with multiple sclerosis (MS), we evaluated free kappa and lambda light chains in the CSF from 3 patients groups: (a) 42 with MS, (b) 16 with other neurological inflammatory diseases (ONID) and (c) 42 with non inflammatory neurological diseases (NIND) used as control. The kappa and lambda light chains contents were evaluated using a specific and sensitive (0.5 mcg/ml) enzyme linked immunosorbent assay (ELISA). Whole CSF immunoglobulins G (CSF-IgG) concentrations were determined by an immuno-nephelometric method and CSF oligoclonal banding was assessed by agarose gel electrophoresis. Elevated free kappa light chains levels were found in 36 of the 42 (85.7%) MS-CSF, and only in 2 of the 16 (12.5%) ONID-CSF. Moreover, all MS-CSF with oligoclonal banding (21/42) exhibited detectable free kappa light chains. In contrast, elevated free lambda light chains or whole CSF-IgG concentrations were found not so specific. These results suggest that free kappa light chains measurement in MS-CSF may have an important diagnostic usefulness. This assay offers a reliable alternative to other available procedures and may constitute complementary technique in routine investigation of MS-CSF.

Cells expressing intracytoplasmic immunoglobulins in secreting and hyposecreting cases of alpha chain disease.

Alpha chain disease proteins (ACDP) originated probably from secreting plasma-cells predominantly present in diffuse and massive enteromesenteric lymphoid infiltration. Very decreased levels of abnormal alpha chain molecules were detected in sera of patients with immunoblastic lymphoma occurring in the late course of the disease. A direct correlation might exist between the proportion of cells bearing intracytoplasmic IgA determinants and the serum amounts of alpha chain disease protein. Relevant evidence raised from study of proliferating lymphoid cells using the unlabeled peroxidase anti-peroxidase method of immunocytochemistry. The percentage of cells expressing intracytoplasmic alpha chains was found to be greater readily secreting case than in hyposecreting case of alpha chain disease. Furthermore, the cells from secreting situation exhibited much more pronounced specific staining, indicative of probably more active synthesis state. Taken together with histological data, these results suggested a possible late evolutionary pathway without detectable intracytoplasmic and serum alpha chain disease protein. They might also support the hypothesis that alpha chain disease and mediterranean lymphoma were different evolutionary phases of the same entity.

Spontaneous enzymatic cleavage of IgD myeloma protein giving a pattern of delta heavy chain disease.

The monoclonality of myeloma proteins is usually demonstrated by their electrophoretic homogeneity and their reactivity with monovalent antisera directed against isotypic determinants of a single heavy chain and a single type of light chain. The absence of precipitation with anti-sera to immunoglobulin kappa and Lambda light chains is a constant character of heavy Chain Disease Proteins (HCDP). However, homogeneous M-components present in the sera of some patients and reacting only with anti-heavy-chain antisera were identified as IgA and IgD myeloma proteins bearing unreactive Lambda chains. In this study, the electrophoretic pattern of a patient serum showed a paraprotein with heterogeneous electrophoretic mobility and precipitation reaction limited to anti-IgD antiserum. The failure to react with anti-light chain antisera was observed by immuno-electrophoresis, immunofixation and rocket-immunoselection. Further analysis by crossed-immunoelectrophoresis revealed that IgD paraprotein contained two separate populations of molecules, one of them being retained when anti-Kappa and Lambda light chains anti-bodies were incorporated in the first dimension gel. It soon became obvious that the observed pattern was generated by enzymatic cleavage of native IgD myeloma protein.

Biclonal gammopathy of kappa light chain type in a case of multiple myeloma.

This immunoglobulin abnormality was found in the serum of a 33 years old man presenting with anemia, weight loss, bone pain and a single bone lesion. The cellulose acetate electrophoresis of his serum showed 2 homogeneous bands migrating in the gamma and alpha-2 regions. Agar gel immunoelectrophoresis using anti-whole normal serum revealed 2 abnormal precipitin arcs of gamma-2 and alpha-2 respective mobility. These 2 M-components precipitated with anti-human kappa light chain antiserum but failed to react with lambda-light chain specific and H-chains specific anti-sera. Furthermore, the 2 monoclonal kappa chains were still demonstrated in the supernatant of the se-rum treated with 50% saturated ammonium sulphate. The 2 kappa chains were isolated from the supernatant using goat anti-kappa antibodies bound to Sepharose-4B. Their SDS-polyacrylamide gel electrophoresis pattern showed a single protein band of 25 Kd molecular weight. This suggest the presence of 2 monomeric kappa chains of different clonal origin. Subsequently, the difference in their electrophoretic mobility was probably due to different V-kappa sequences. However, it is also possible that the unusual alpha-2 mobility is a consequence of a moderate glycosylation without increase in molecular weight.

[Rapid and sensitive micromethod for protein determination by the Coomassie-blue technique]. / Micro-methode rapide et sensible de dosage des proteines par la technique au bleu de Coomassie.

The Coomassie blue method of proteins quantitation initially reported by Bradford and modified by Macart et al. appears to be simple, fast, sensitive and of low cost. In this study, we report an improved micromethod derived from the technic described by Macart. Applied to the protein quantitation of 100 cerebrospinal fluid (CSF) specimens and 50 serum specimens our method showed a very good correlation with the macromethod of Macart (r = 0.985 for CSF proteins, r = 0.944 for serum proteins) and also with the Lowry technic (r = 0.986 for CSF proteins, r = 0.898 for serum proteins). Our technic may be used with various biological fluids (urines, tears, saliva...) and seems to be of particular interest for low protein level fluids and for little volume samples (it needs only 20 mcl).

[Detection of oligoclonal immunoglobulins in the cerebrospinal fluid using agar gel immunoelectrophoresis with silver salt staining]. / Détection des immunoglobulines oligoclonales dans le l. c. r. par IEF sur gel d'agarose et coloration par sel d'argent.

Detection of oligoclonal immunoglobulins in multiple sclerosis cerebrospinal fluid (MS-CSF) seems to be an important test in the biological diagnosis of the disease. In our laboratory, the classical agarose gel electrophoretic technic allowed the detection of CSF oligoclonal bands in only 40% of the MS patients. This relatively low percentage in comparison with those obtained by other investigators led us to develop a much more resolving electrophoretic technic: an agarose gel isoelectric focusing (IEF) with silver staining. By this method, we detected oligoclonal immunoglobulins in 43 (43%) of 100 MS-CSF exhibiting only polyclonal patterns on classical electrophoresis. Oligoclonal banding appeared particularly in patients with inflammatory type profile (36/60) according to Schuller's classification, but also in those with normal (5/26) or inflammatory transudative (2/8) type profiles. MS-CSF with oligoclonal immunoglobulins on IEF had a higher IgG/Albumin ratio (p = 0.01) than those with polyclonal immunoglobulins, while the mean IgG levels were not significantly different (p = 0.07). These results support the diagnostic usefulness of the IgG/Albumin ratio. Agarose IEF appears to be a useful technic in the detection of oligoclonal immunoglobulins and may be applied at least to CSF of patients with clinical signs of multiple sclerosis.

[Immune response to tetanus toxoid in mice receiving protein deficient diet immediately after weaning]. / Etude de la réponse immunitaire au vaccin antitétanique chez des souris soumises, au moment du sevrage, a une carence alimentaire protéique.

The humoral immune response to tetanus toxoid is evaluated by passive haemagglutination test in mice receiving low diet immediately after weaning during 15 and 30 days. The results show that the more deficient mice give the best antitoxin titers but after the challenge the antibody immune response become proportional to the protein restriction. In other respects and antitoxin titers are higher in 30 days restricted mice than in 15 days deficient ones. The effect of Bordetella pertussis adjuvant on the immune response of protein deficient mice seems nul. These results are discussed.

[Comparison of reversed phase passive hemagglutination test and the electro-immunodiffusion technic for the detection of HBs antigen]. / Etude comparée de l'hémagglutination passive inversée et de l'électro-immunodiffusion dans le dépistage de l'antigène HBs.

In this study, were compared two methods used in case-finding of the australian antigen (Ag. HBs) : the electro-immunodiffusion technique (E.I.D.) and the passive reverse hemagglutination tests. All the tests were carried out on couple of sera obtained from mothers and her newborns, giving on the whole 717 sera. The results we obtained led us to maintain the E.I.D. for the biological diagnosis of B virus hepatitis. The passive reverse hemagglutination test has been kept back for the titration of the Ag. HBs in the sera found positive by E.I.D. With regard to the vertical transmission (mother-newborn) of Ag. HBs, we could not draw any good conclusion.