Search

A chromatographic method was employed to study the kinetics of human serum albumin (HSA) adsorbed on immobilized monoclonal antibodies. The antibodies of various specificities were covalently bound to a high-performance liquid chromatography (HPLC) silica support. For very low desorption rates, successive amounts of the reacting protein were injected until column saturation. The analysis of the increase of the non-retained fraction calculated from peak area measurements gives the capacity of the support and the rate of the biospecific adsorption process. The model is based on a second-order Langmuir kinetic law and assumes a global mass transfer for the adsorption process. The use of a silica support of small pore size permits the reduction of the contribution for mass transfer in the stagnant fluid and the decrease in the column capacity: due to its large size, the reacting molecule is adsorbed on the external surface of the particle. The adsorption rate constants of HSA on five monoclonal anti-HSA antibodies of different specificities were determined. For all the immuno-adsorbents studied, the adsorption rate constant is significantly lower than that found on immobilized polyclonal antibodies. Measurements at different flow rates reveal that the mass transfer due to the transport to the adsorbent surface is small and can be estimated.

The specificity of six peroxidase-labelled anti-albumin monoclonal antibodies was studied by allowing them to react with albumin-derived fragments of known structure comprising the whole albumin molecule. The epitopes corresponding to these antibodies were located on defined regions of the albumin molecule extending from the N to the C-terminal ends. These antibodies have been used for investigating by ELISA the conformational alterations of albumin molecule brought about by the N-B transition in twelve individual sera. The results do not show any significant differences between the sera. The N-B transition involves essentially the N-terminal portion of the albumin molecule. There also exists as modification of the C-terminal region which is however much less pronounced than the one in the N-terminal region. In addition, it is not observed when using isolated albumin.

A 75 year old man developed chronic flexor tenosynovitis in a finger, caused by Mycobacterium terrae. He had a history of diabetes mellitus and arteritis, but not of local trauma. However, triamcinolone had been injected into the tendon sheath on 3 occasions. Chemotherapy was instituted, but after 3 months amputation became mandatory. This is the fourth case reported in which M. terrae was involved, and it supports the responsibility of this organism as a human pathogen. Attention is drawn to the hazards of corticosteroid injections into tendon sheaths.

Fragments A, B and C obtained by CNBr degradation of human serum albumin (HSA) were studied by specific quantitative precipitation with anti-HSA sera. A co-precipitation has been observed between fragments B and C as well as between C and A which follow each others in the albumin molecule. On the contrary, there was no co-precipitation between fragments B and A which correspond respectively to the N- and C-terminal part of the albumin molecule. Moreover, using inhibition of passive haemagglutination, it has been observed that fragment A does not inhibit anti-B antibodies and fragment B does not inhibit anti-A antibodies. These results indicate that there are common antigenic sites to fragments B and C as well as to C and A but not to B and A. This is in agreement with the data upon the structure of HSA showing that it is made of three domains in linear sequence.

Twelve recent cases of Plasmodium falciparum malaria presented with unusual clinical and laboratory features. All patients had regularly been taking adequate doses of amino-4-quinolines as prophylaxis. In most cases the symptoms were mild as compared with those in a group of 20 control patients with typical malaria. More surprisingly, parasitaemia was either negative or very weakly positive (less than 1000 infested erythrocytes per mm3). All cases occurred in people returning from areas in which resistance of P. falciparum to these drugs is endemic. Diagnostic problems could be solved in most patients by new immunological techniques relying on the detection of parasitized erythrocytes and plasmodium antigens using a battery of monoclonal antibodies. A physiopathological model explaining why parasitaemia is negative in such cases is suggested.

An enzyme-linked immunosorbent assay for measuring penicillin groups is described. Alkaline phosphatase conjugated to penicilloyl residue was reacted with purified anti-penicilloyl antibodies coated upon polystyrene plates. The inhibition of this reaction allowed a specific determination of penicilloyl residues at the picomole level. This assay was applied to the study of the penicilloyl groups linked to human albumin and to the measurement of some of these groups after enzymatic degradation or physical modifications of the albumin molecule.

The antigenic structure of a fragment (F1) of human serum albumin (HSA), carrying one of the antigenic sites of the whole molecule, has been studied. Previous studies have shown that this fragment is made of two peptide chains linked by a disulfide bond. One of these chains named alpha has 28 residues and one intrachain disulfide bond giving rise to a loop of 9 or 10 residues. The chain occurs under two forms: the beta chain made of 25 residues and the gamma chain which differs from the beta chain by the absence of the two last C-terminal residues (F. Bellon and C. Lapresle, Biochem. J., 1975, 147, 585-592). The alpha and beta chains both inhibit independantly at 100% the agglutination by an anti-albumin serum of red cells sensitized with F1. Chains alpha, beta and gamma displace anti-HSA antibodies fixed upon an immunoadsorbent prepared with F1. Chain alpha is more active than chain beta which in turn is more active than chain gamma. Reduction and alkylation of alpha and beta chains abolish almost completely their immunological activity. Immunoadsorbents prepared with alpha or beta chains adsorb all of the anti-F1 antibodies from an anti-HSA serum. Antibodies isolated from each chain are inhibited by both chains. Chains alpha and beta induce in rabbits the formation of antibodies reacting with F1 and HSA, alpha chain being a better immunogen than beta chain.

Between April and June 1984, eight chloroquine-resistant P. falciparum malaria cases were observed in non-immune patients after their emergency sanitary return from NE Mozambique. In vivo resistance to amodiaquine and pyrimethamine were demonstrated from two isolates. Radical cure was obtained with mefloquine in all cases. These patients came from an expatriate community of 35 adult men among which six other chloroquine-resistant malarial cases were probable. The epidemic pattern of this chloroquine-resistant focus is underlined.

We have studied the reaction of two monoclonal anti-human albumin antibodies with various fragments of albumin comprising the entire molecule. Using solid phase assay, inhibition of enzyme immunoassay and of passive hemagglutination, they were shown to be specific for a fragment F1, of 6000 dalton, located near the C terminus of human albumin. In addition, these antibodies reacted weakly with fragment D which corresponds to the N terminal half of the molecule.

The authors report the case of an eight-year-old boy who developed erythroderma under carbamazepine. Other findings were enlarged lymph nodes, slight hepatic cytolysis, leuconeutropenia and decrease in the three classes of immunoglobulins. The immunosuppressive effect of the drug is recalled. Respective responsibility of immunosuppression and allergy is discussed.

Two fixation sites for penicilloyl groups on human albumin were demonstrated. Using CNBr cleavage the first site was located between methionine 123 and methionine 297 and the second one between methionine 297 and the C-terminal residue. In both cases, penicilloyl groups were unmasked by pronase degradation or disulfide bond reduction.

A comparative analysis of subsets (4B4 and 2H4) within the CD4 lymphocyte subpopulation was made by double marker analysis among 23 healthy heterosexual, 16 healthy HIV seronegative high-risk homosexuals and 82 HIV seropositive subjects. Data show that the significant increase in CD4 lymphocytes observed among seronegative homosexuals corresponds mainly to an increase in CD4-4B4 positive cells while CD4-2H4 subset levels remain comparable to healthy heterosexual controls. A decrease both in CD4-4B4 and CD4-2H4, parallel to a decrease in CD4 subpopulation, was observed in asymptomatic seropositive carriers (SPC) and patients with ARC syndrome (AIDS-related complex) or AIDS (acquired immunodeficiency syndrome). Interestingly, an absolute decrease in CD4 subpopulation in patients with asymptomatic lymph node enlargement (LAS), is chiefly accounted for by a decrease in CD4-4B4 subset. The values of CD4-2H4 subset are significantly higher than those observed for SPC patients and are close to the normal values. These observations are of interest, as no differences between SPC and LAS patients could be detected when CD3, CD4 and CD8 subpopulations were studied.

The case described is one of endocarditis due to lactobacillus casei in a 33-year-old man with aortic insufficiency of undetermined origin. The clinical signs were neurological, following thrombosis of the right posterior cerebral artery. The source of injection was a dental extraction. This case, with a typical clinical picture, is the eleventh to be reported in the literature.

Five proteins (albumin, orosomucoid, IgG, IgM and IgA) were assayed by simple radial immunodiffusion method in the sera of 345 healthy subjects and 5 000 patients suffering from some 200 different diseases. The results were analyzed by computer, and from concomitant changes in individual protein levels it was possible to divide the subjects into eleven groups, each group having a distinctive protein profile. These profiles are characteristic of either a disease (e.g. group VI for virus hepatitis, group VII for trypanosomiasis) or of a physiopathological process (e.g. group II for inflammation, group V for liver damage, group VIII for hypogammaglobulinaemia due to anabolic deficiency and group IX for the same condition due to passage of serum proteins into the extravascular compartment), or they have a prognostic value (e.g. group I).

When human albumin was treated with CNBr, a fragment designated D was obtained and attributed to the absence from some of the albumin molecules of methionine at position 123 [Lapresle & Doyen (1975) Biochem. J. 151, 637-643]. The present study shows that methionine-123 is converted into homoserine without cleavage of the subsequent methionine-cystine bond. With bovine alpha-lactalbumin, a further example of non-cleavage of a methionine-cystine bond with conversion of methionine into homoserine is reported.

Analysis of the antigenic structure of human serum albumin was undertaken using monoclonal antibodies. Nineteen antibodies were prepared and their specificities were studied using fragments which encompass the whole sequence of the albumin molecule. These antibodies recognized 13 different epitopes which are different from the one previously identified with two other monoclonal antibodies [Doyen et al., Immun. Lett. 3, 365-370 (1981)]. Among those 13 different epitopes, six were overlapping. Four epitopes were located on the N-terminal half of the albumin molecule. One of these required integrity of methionine 87 and the other three were overlapping and located around methionine 123. Eight epitopes were located on the C-terminal half of the albumin. Two of them were within the sequence, 330-422 and 299-496 respectively; the other six appeared to be topographic determinants which were altered or lost in the albumin fragments. A last epitope could not be located on any region of albumin. Four monoclonal antibodies directed against a given portion of the albumin molecule reacted slightly with another part of albumin, thus confirming the existence of an intramolecular cross-reactivity between the different domains of human albumin.

A method for studying conformational changes induced in the human albumin molecule, either in its purified form or in serum, is described. Plates were coated with albumin or human serum at varying pHs and were reacted with peroxidase-labeled anti-albumin monoclonal antibodies of different specificities. The data showed that albumin molecules were coated in conformations induced as a result of pH changes, allowing us to demonstrate that pH modifications involved the N-terminal portion of the albumin molecule whether in its purified form or in serum. This method should be applicable to the study of conformational modifications in other proteins as well.

The high frequency of meningococcal infections in patients with congenital deficiency of a component of the complement terminal pathway emphasizes the critical role of this system in host resistance against Neisseriaceae. We report the observation of a subacute septicaemia due to N. meningitidis serogroup Y. This girl had an acquired deficit of the C3 fraction of complement due to a high titre of C3 nephritic factor (C3NeF). There was no evidence of partial lipodystrophy or biological symptoms of glomerular disease. The meningococcal infection revealed this biological abnormality.

In a 26-year-old man who had spent time in a trypanosome endemic zone, lesions of erythema marginatum progressing by acute exacerbations for 2 years despite various forms of treatment led to the discovery of a multiple lymphadenopathy and a greatly accelerated sedimentation rate. These findings suggested a diagnosis which was confirmed by the discovery of Trypanosoma gambiense in the blood and lymph node aspirate. In addition, there was an albumino-lymphocytic reaction in the cerebrospinal fluid. Finally, an atrioventricular block was discovered. All these symptoms and signs responsed to treatment with melarsoprol but a pericardial reaction developed, possibly of immunoallergic origin.

Two monoclonal anti-HSA antibodies, HA1 and HA2, have been shown to be specific for a univalent fragment of 6000 mol. wt, F1, located near the C-terminus of HSA (Doven, Pesce & Lapresle, Immunology Letters 3, 365-370, 1981). Both monoclonal antibodies have been shown to react with the same site, which includes the following components: the last small loop of HSA (558-567) the disulfide bridge 514-559 and the residue 570. This site is as available on HSA and F1, but partially masked on the 'Inhibitor' fragment from which F1 derives. Polyclonal anti-F1 antibodies, purified from rabbit sera or mouse ascites by affinity chromatography, react with the same site as HA1 and HA2. However, polyclonal antibodies are heterogenous, most probably because they consist of anti-F1 specific antibodies and of antibodies specific against other parts of the albumin molecule which cross-react with F1. In addition, monoclonal antibodies can recognize the mutation of a single amino acid residue in the albumin molecule.