A combined indirect ELISA and immunoblotting for the detection of intrathecal herpes simplex virus IgG antibody synthesis in patients with herpes simplex virus encephalitis.

A combined indirect ELISA and immunoblotting assay was used for the detection of intrathecal synthesis of IgG antibodies to herpes simplex virus (HSV) in patients with HSV encephalitis (HSVE). By using these two assays as well as three markers for blood-brain barrier, leakage can be easily excluded. A total of 21 sera and 24 cerebrospinal fluid (CSF) samples from 11 patients with HSVE were examined. For seven patients more than one pair of serum and CSF were available. For one patient IgG antibodies began to be detectable in CSF after the sixth day from the onset of the disease. In the other 10 patients the intrathecal synthesis of HSV IgG antibodies was detected later than the sixth day and reached high optical density (OD) values after the 10th day from the onset of disease, at the earliest. In contrast, intrathecal HSV antibody synthesis was not found in specimens taken from 20 patients with acute meningitis who composed our negative control group. The use of a combined indirect ELISA and of an immunoblotting assay on a single dilution of serum and CSF for HSV IgG synthesis in the central nervous system (CNS) allowed the diagnosis of HSVE after the first week of disease.

Herpes simplex virus types 1 and 2 and cytomegalovirus in peptic ulcer disease and non-ulcer dyspepsia.

The prevalence and the serum levels of IgG antibody to Herpes simplex virus type 1 or 2 (HSV1, HSV2) and to cytomegalovirus (CMV) were studied by ELISA in patients with active peptic ulcer -- duodenal and gastric -- and non-ulcer dyspepsia. Two hundred and forty-two consecutive patients with endoscopically confirmed active peptide ulcer -- 170 duodenal ulcers, 72 gastric ulcers -- and 95 consecutive patients who fulfilled the criteria for the diagnosis of non-ulcer dyspepsia were included in the study. The patients, aged 17-80 years, were well matched for age and sex. Antibody to cytomegalovirus was found in 83% of duodenal ulcer, 85% of gastric ulcer and 75% of non-ulcer dyspepsia patients; differences were not significant. The prevalence of HSV1 antibody was significantly higher in patients with duodenal ulcer than in those with non-ulcer dyspepsia (p < 0.025); the prevalence of HSV2 antibody was significantly higher in patients with duodenal or gastric ulcer, than in those with non-ulcer dyspepsia (p < 0.05, p < 0.01, respectively); however, antibody levels (mean optical density) to the viruses studied were similar for all groups of patients. These results provide some evidence that HSV might be implicated in the pathogenesis of peptic ulcer disease.

The main immunogenic region of the acetylcholine receptor. Structure and role in myasthenia gravis.

Auto-antibodies to the nicotine acetylcholine receptor (AChR) cause the disease myasthenia gravis (MG). Animals immunized with AChR or receiving anti-AChR antibodies acquire MG symptoms. The majority of the monoclonal antibodies (mAbs) raised in rats against intact AChR bind to a region on the extracellular side of the AChR's alpha-subunit, the main immunogenic region (MIR). The major loop of the overlapping epitopes for several anti-MIR mAbs has been localised between residues 67-76 of the alpha-subunit. Anti-MIR mAbs are very potent in accelerating AChR degradation (antigenic modulation) in muscle cell cultures and transferring experimental MG in animals. Fab fragments of single anti-MIR mAbs when bound to the AChR inhibit two-thirds of the MG patients' antibodies from binding and from inducing antigenic modulation of the AChR. This suggest that the majority of the human MG antibodies are also directed against the MIR. It has however to be verified by direct experiments.

Monoclonal antibodies against the main immunogenic region of the acetylcholine receptor. Mapping on the intact molecule.

About two-thirds of the antibodies to the nicotinic acetylcholine receptor (AChR) in myasthenic patients, and in rats immunized with intact AChR, bind to the main immunogenic region (MIR) on the alpha-subunit. We tested all available anti-MIR monoclonal antibodies (mAbs) by competition experiments for binding on the intact AChR from Torpedo electric organ and human muscle. Practically complete competition between all possible paired combinations of anti-MIR mAbs was found. As a consequence, the MIR must be a very concrete and small region. Furthermore, the location of the MIR relative to some other less immunogenic regions was also determined.

Conformation of cytoplasmic segments of acetylcholine receptor alpha- and beta-subunits probed by monoclonal antibodies: sensitivity of the antibody competition approach.

The conformation of the cytoplasmic side of Torpedo marmorata acetylcholine receptor (AChR) was investigated by 22 monoclonal antibodies (mAbs) binding to known sites on the amino acid sequences 339-378 and 336-469 of the AChR alpha- and beta-subunits respectively. Competitions among these mAbs for binding on the intact AChR were compared with their competition for binding on the SDS-denatured subunits and with their corresponding epitopes previously determined on the primary structure of the subunits. We found the following: The three approaches correlated very well suggesting that these mAbs bind on the intact AChR at the same sequences determined by synthetic peptides and not on irrelevant discontinuous epitopes; this finding supports conclusions of Ratnam et al. (1986a) that the amphipathic helix M5 is exposed on the cytoplasmic side of the AChR. The subunit segments alpha 339-378 and beta 336-469 seem to be extended over large distances on the cytoplasmic surface of the AChR. The cytoplasmic surface of beta-subunit has a very immunogenic region. The mAb-competition technique is very sensitive since mAbs to epitopes separated by only about seven residues did not exclude each other, and mAbs to overlapping epitopes exhibited differential competitions with other mAbs.