Impaired parvocellular pathway in dyslexic children.

Recent studies report that some children with dyslexia have impaired visual processing, specifically in the fast-processing magnocellular pathway. The objective was to study the effect of varying luminance and temporal and spatial frequency on the latency and amplitude of the visual evoked potentials (VEPs) in normal and dyslexic Egyptian children who speak Arabic (a right-left reading and writing system). VEPs were recorded in 52 dyslexic and 41 normal children in the fourth grade using a black and white checkerboard pattern with different checkerboard sizes and different rates of stimuli at high- and low-contrast media. The peak of the major positive wave component (P100) of each waveform and the trough of the previous major negative wave component were identified, and the peak-to-trough amplitude was measured. The latency and amplitude of VEPs in response to different experimental conditions showed significant shortening of P100 latency under high-contrast media and under low spatial frequency in children with dyslexia compared with normal readers. Furthermore, dyslexia children showed prolonged P100 latency in response to high spatial frequency stimulation compared with the low spatial frequency (P=0.003) and significantly higher N1-P1 amplitude under high-contrast media compared with low-contrast media (P=0.02), whilst no such changes were observed in normal readers. These results are suggestive of deficiency within the parvocellular pathway rather than the magnocellular pathway. As reading apparently places demands primarily on the ability to discriminate fine details, which is to say, on the parvocellular system, we suggested that deficiency in this system, at least in Arabic speaking children, could be a predisposing factor in dyslexia.

Cerebrospinal fluid levels of amyloid precursor protein and amyloid beta-peptide in Alzheimer's disease and major depression - inverse correlation with dementia severity.

Alzheimer's disease (AD) is the most common neurodegenerative disorder characterized by progressive dementia that ultimately leads to death. Histopathological hallmarks of AD include brain amyloid deposits and neurofibrillary tangles. Major depression is a frequent diagnosis in every gerontopsychiatric clinic that sees patients with both cognitive and affective disorders. Many depressed patients, in fact, are clinically characterized by cognitive impairments. Thus, an assay that excludes - or confirms - probable AD in cognitively impaired patients is desirable. Such assays may use protein markers that are derived from such histopathologically relevant molecules as the amyloid precursor protein (APP) and its derivatives including the amyloid beta-peptides (Abeta). To evaluate the differential diagnostic properties of cerebrospinal fluid (CSF) Abeta and secreted soluble ectodomain (APPs), we quantitated CSF levels of these measures in AD patients and compared them to age-matched control patients with major depression. CSF levels of APPs and Abeta were similar in patients with AD or major depression, and the apolipoprotein E genotype had no influence on CSF levels of Abeta in AD patients. Measurement of Abeta peptide using a novel zinc/copper capture ELISA that detects aggregated Abeta peptides as well demonstrated similar levels in AD and major depression. In AD patients, CSF levels of total Abeta (Abeta1-40 plus Abeta1-42) were inversely correlated with a functional measure of dementia severity (NOSGER), suggesting that CSF levels of Abeta decrease with advancing severity of AD. Thus, CSF levels of Abeta are not useful for the differentiation of AD from major depression. However, CSF levels of Abeta reflect the severity of dementia and may be useful as biological markers of the stage of the disease.

A new immunoassay for soluble fibrin enables a more sensitive detection of the activation state of blood coagulation in vivo.

A novel sandwich immunoassay for measurement of soluble fibrin in plasma has been developed. For immunization we used the synthetic heptapeptide Gly-Pro-Arg-Val-Val-Glu-Arg representing the amino terminus of the alpha-chain of human fibrin. A monoclonal IgG1 antibody was obtained by conventional hybridoma technology. To increase convenience, the sandwich immunoassay was developed for the Enzymun-Test systems which are based on streptavidin pre-coated tubes. The new assay was designed with the same fibrin specific antibody both in biotinylated and in peroxidase-labelled form. Fibrin in native plasma samples could only be detected after pre-incubation of the plasma with chaotropic ions. Test results were calculated using a standard curve comprising six fibrin standards (0-50 micrograms/ml). Precision of the method was satisfactory; intra-assay CVs using plasma samples ranged between 5.0% (23.7 micrograms/ml) and 12.4% (0.2 microgram/ml). CVs of interassay precision measurements using standards as samples range between 7.3% (25.0 micrograms/ml) and 11.4% (1.0 micrograms/ml). The lower detection limit was 0.12 microgram/ml. Investigations of normal range in 70 age-matched healthy individuals resulted in a mean of 1.12 micrograms/ml. Linearity was excellent; recovery of high fibrin plasma after dilution with normal plasma was always between 100 and 107%. Fibrin specificity was due to the monoclonal antibody 2B5 used and no cross reactivity with fibrinogen, fibrinogen split products or fibrin D-dimer was observed. Fibrin fragment E1 (studied by Dempfle CE, et al. Blood Coag Fibrinol 1993; 4: 79-86) and fragments X and Y showed moderate cross reactivity in the assay and caused some overestimation of fibrin at high fibrin split product concentrations.(ABSTRACT TRUNCATED AT 250 WORDS)

[Enzyme immunoassay for the detection of autoantibodies to nRNP and Sm: a rapid and sensitive alternative to current procedures]. / Enzymimmunoassays zum Nachweis von Autoantikörpern gegen nRNP und Sm: eine schnelle und sensitive Alternative zu den bisherigen Nachweisverfahren.

Antinuclear antibodies are of major importance in the diagnosis of inflammatory rheumatic diseases. Sm and nRNP antibodies can be found in sera from patients with systemic lupus erythematosus and mixed connective tissue disease. Usually, these antibodies have been detected with one of the following methods: Ouchterlony immunodiffusion, passive hemagglutination or counterimmunoelectrophoresis (CIE). In this work results obtained by Ouchterlony and CIE techniques were compared with those obtained by ELISA. Purified proteins from cellular extracts (HeLa) were used as antigens for Sm- and nRNP-ELISA: D polypeptide for Sm-ELISA and the 68 kD, A, C, B,B' and D polypeptides for nRNP-ELISA. Compared with the other two techniques, ELISA was less time consuming and showed greater sensitivity. Quantitative titration proved to be of advantage in monitoring the course of the diseases mentioned above.

Single incubation multilayer immune technique.

A new method termed the 'single incubation multilayer immune technique (SIMIT)' is described which features two striking advantages over standard immunoassay techniques: shorter assay time and superior assay sensitivity. Using standardized, optimized amounts of ligand (e.g., antibodies) and anti-ligand (e.g., anti-antibody) it is possible to co-incubate these two reactants so that during a single incubation step multiple layers of immunoreactants are formed thereby resulting in enhanced assay sensitivity. Depending upon the application, SIMIT improves detection levels by about 10-20-fold. This report describes the use of SIMIT for two completely different assay systems: (a) indirect enzyme-linked immunosorbent assays, and (b) unlabeled enzyme immunoassays. However, SIMIT should be applicable to a large variety of other assay systems.

Quantitation of the anaphylatoxin C3a in the presence of C3 by a novel sandwich ELISA using monoclonal antibody to a C3a neoepitope.

C3a levels in plasma are usually measured by a competitive inhibition radioimmunoassay (RIA) using 125I-labelled C3a-desArg and antibodies to C3a capable of detecting C3a determinants which are also present on the native C3. Therefore, prior to the assay native, non-cleaved C3 has to be removed completely from the C3a-containing sample by precipitation. We developed a new rapid two-site sandwich ELISA system for the quantitation of C3a-desArg in plasma. This immunoassay uses a monoclonal antibody (mAb H466) reacting with C3a-desArg but not with C3. The reactivity of mAb H466 with a neoantigenic determinant of C3a-desArg permitted the direct quantitation of C3a-desArg without removal of C3 from the sample. The mAb H466 was used as a capture antibody and bound C3a-desArg was detected with a second peroxidase-labelled anti-C3a mAb. The lower limit of detection of C3a-desArg in this ELISA was 1 ng/ml. The C3a-desArg levels measured in the plasma samples of various patients were found to differ over a wide range. A good correlation was observed between the results obtained in the RIA and those obtained in the ELISA (r = 0.95). High levels of C3a-desArg were detected in plasma from patients with multiple trauma and patients undergoing haemodialysis. The C3a-desArg assay described should facilitate the routine quantitation of C3a in samples of plasma.

The C terminus of the anaphylatoxin C3a generated upon complement activation represents a neoantigenic determinant with diagnostic potential.

Activation of the C component C3 results in generation of the anaphylatoxin C3a. The C3a polypeptide chain consists of 77 amino acids. The active site of this potent mediator, which also has immunoregulatory function resides in its C terminus. This report demonstrates that the C terminus of C3a (C3a-desArg) exposed by proteolytic cleavage from C3 represents a neoantigenic determinant. Two mAb specific for this epitope were obtained after immunization with the synthetic octapeptide (OP) Arg-Ala-Ser-His-Leu-Gly-Leu-Ala [C3a(69-76)] coupled to the carrier keyhole limpet hemocyanin (KLH). These anti-C3a(69-76) antibodies (H453 and H454) reacted in an ELISA system with C3a and KLH-OP but not with C3 or with KLH alone. Free OP efficiently blocked binding of the antibodies to C3a, whereas binding of another anti-C3a mAb (H13) remained unaffected. In immunoblotting analysis, the anti-C3a(69-76) mAb reacted with purified C3a but failed to react with the denatured, noncleaved C3. A novel quantitative C3a-ELISA was established with the anti-C3a(69-76) mAb. It had a sensitivity in the nanogram range (1 to 5 ng/ml). The C3a determination was not impaired by the presence of high concentrations of C3. Therefore, C3 removal was not required in contrast to the previously described C3a assays. This C3a ELISA might facilitate clinical C3a quantitation, e.g., in samples from patients with adult respiratory distress syndrome. In these patients, C3a determination in the early phase of the disease is of diagnostic relevance and has prognostic value.

The cellular substrate: a very important requirement for baculovirus in vitro replication.

We established more than 200 primary cell lines of Cydia pomonella (coding moth). 81 of them were selected and screened for replication of two baculoviruses (from two different subgroups): the Choristoneura murinana NPV and the Cydia pomonella GV. Although all these cell lines had been derived from the same insect species, they varied largely in their response to challenge with the NPV. Most of them showed CPE or produced different amounts of polyhedra. Interestingly, we also found a few cell lines that were permissive for GV replication. To our knowledge this is the first time that GV replication in cell lines has been obtained. Our results show that cell line properties are most important for baculovirus in vitro replication.

Characterization of monoclonal antibodies to the Autographa californica nuclear polyhedrosis virus.

Mouse hybridomas which produce monoclonal antibodies to the Autographa californica nuclear polyhedrosis virus (AcNPV) have been prepared and the protein to which each is directed has been determined. The antibody produced by clones 3F3 and 5E9 specifically bound polyhedron protein of MW 32,000 when used in immunoadsorption chromatography When used in ELISA they could not distinguish between insect- and cell culture-derived polyhedron protein and also reacted with a wide range of baculoviruses. The antibody produced by clone 3D10 specifically immunoprecipitated a polypeptide of MW 42,000 from tissue culture-derived extracellular virus which was a prominent product synthesized in infected cells. This polypeptide was a major component of enveloped virus, released from insect-derived polyhedra, and nucleocapsids. In contrast to 3F3 and 5E9, this antibody was specific for AcNPV.