A study to determine plasma antioxidant concentrations in patients with Barrett's oesophagus.

BACKGROUND: Dietary questionnaire studies have suggested that patients with oesophageal adenocarcinoma are deficient in antioxidants. It is not known whether the same holds true for patients with the precursor lesion, Barrett's oesophagus. AIMS: To evaluate the hypothesis that patients with Barrett's oesophagus are deficient in antioxidants compared with patients without evidence of Barrett's oesophagus. PATIENTS AND METHODS: Plasma antioxidant profiles (copper, selenium, zinc; vitamins A, C, and E; carotenoids) were determined for patients with Barrett's oesophagus (n = 36), patients with erosive oesophagitis (n = 32), and patient controls (n = 35). RESULTS: Patients with Barrett's oesophagus had significantly lower plasma concentrations of selenium, vitamin C, beta cryptoxanthine, and xanthophyll compared with the other groups. CONCLUSIONS: This study confirms the hypothesis that patients with Barrett's oesophagus are deficient in certain antioxidants.

The prevalence of anti-acetylcholinesterase antibodies in autoimmune disease.

A robust and precise enzyme linked immunosorbent assay (ELISA) with proven sensitivity and specificity has been employed to detect human antibodies (allogenic/autogenic) to human acetylcholinesterase (AChE). The sensitivity of the method has been established using mouse monoclonal antibodies (0.8 ng/ml) and uniquely, human sera positive for anti-Yt(a) allogenic antibodies, to one phenotypic form (most common) of human AChE. The latter was also used as the positive human control to ensure functionality of the assay. The ELISA method was used to establish a normal distribution curve for absorbance values employing sera from healthy blood donors Subsequently, the ELISA was employed to investigate the prevalence of anti-AChE antibodies in patients with confirmed autoimmune disease and patients with non-autoimmune thyroid disease (diseased control). The results indicate that there is not a high prevalence of anti-AChE antibodies in patients with confirmed autoimmune disease. The lack of anti-AChE autoantibodies in patients' with clinically apparent Graves' ophthalmopathy, mitigates against there being a causal role of such antibodies in Graves' associated eye disease.

A potential role for free radical-mediated skeletal muscle soreness in the pathophysiology of acute mountain sickness.

BACKGROUND: It has been suggested that free radicals may be implicated in the pathophysiology of acute mountain sickness (AMS) due to their ability to initiate and propagate cell membrane damage (3). Therefore, the present study was designed to: a) investigate the effects of an expedition to high altitude on metabolic indices of free radical-mediated oxidative stress and assess subsequent implications for skeletal/cardiac muscle damage; and b) determine whether these parameters were different in subjects who developed AMS after gradual ascent to 5100 m (base camp, BC) compared with those who remained healthy. METHODS: There were 19 male volunteers who were examined at rest and after a standardized maximal exercise test at sea level before and after an expedition (SL1/SL2) and during the first morning of arrival at BC. The trek to BC lasted 20+/-5 d. RESULTS: A mild increase in the Lake Louise AMS score was observed by the end of day 1 at BC (p < 0.05 vs. SL1/SL2). Four subjects developed AMS, which in one subject later progressed to high altitude pulmonary and cerebral edema. The serum concentration of lipid hydroperoxides (LH) increased markedly at rest and after maximal exercise at BC (p < 0.05 vs. SL1/SL2) whereas no changes were observed for plasma malondialdehyde (MDA). Resting serum total phosphocreatine kinase activity (CPK) and myoglobin also increased at BC (p < 0.05 vs. SL1/SL2) whereas cardiac troponin I (cTnI) remained stable. The resting pain threshold decreased and exercise-induced muscle soreness subsequently increased at BC (p < 0.05 vs. SL1/SL2). An association was observed between resting LH and myoglobin at BC (r = 0.45, p < 0.05) and the increase in LH was related to the increase in exercise-induced muscle soreness at BC (r = 0.96, p < 0.05). Further correlations were identified between the AMS score on day 1 at BC and: a) resting/exercise LH (r = 0.63, p < 0.05/r = 0.51, p < 0.05); and b) resting pain threshold at BC (r = -0.58, p < 0.05). Furthermore, subjects with AMS on day 1 at BC were characterized by a greater decrease in the resting pain threshold and greater increase in resting LH, CPK and myoglobin compared with subjects without AMS (p < 0.05). Headache, fatigue, insomnia and general apathy were the most frequently reported symptoms of AMS. CONCLUSIONS: Localized free radical-mediated vascular damage of the blood-brain barrier in addition to systemic tissue damage causing overt skeletal muscle soreness may have contributed to the pathophysiology of AMS, the latter through its indirect effects on other non-specific constitutional symptoms such as fatigue and insomnia causing a deterioration in physical performance.

Autoantibodies to acetylcholinesterase revisited.

A sensitive and specific enzyme linked immunosorbent assay (ELISA) utilizing human recombinant acetylcholinesterase has been employed for the detection of human antibodies to human acetylcholinesterase. The method can detect allogenic antibodies to the Yt(a) form of human erythrocyte AChE. Adaptation of this ELISA method allowed the IgG subclass typing of IgG anti-AChE antibodies, which could help to determine the possible role of these antibodies in the aetiology of any neurological conditions. Routine serological investigations established the AChE phenotype of each of the patients recruited, to determine whether anti-AChE antibodies were allogenic or autogenic in origin. These techniques were used to determine the incidence of autoantibodies to AChE in patients with neurological conditions, including the subtypes of motor neuron disease. The data presented are not consistent with earlier reports of a high incidence of autoantibodies to AChE in amyotrophic lateral sclerosis and progressive muscular atrophy.

Detection of anti-Yta antibodies using a sensitive and specific enzyme-linked immunosorbent assay.

A specific, sensitive and semi-quantitative enzyme-linked immunosorbent assay (ELISA) is described to detect anti-Yta antibodies in human serum. Recombinant acetylcholinesterase (AChE E.C.3.1.1.7) was employed as the coating antigen in the microtitre plate and horseradish peroxidase (HRP)-conjugated specific antibody (IgG) was used as the secondary antibody. The method developed showed excellent sensitivity, detecting a titre > 1 in 600,000 (3.5 ng/mL mouse IgG protein) for mouse monoclonal (mMAb) anti-AChE antibody. No cross-reaction was seen with other common blood group antibodies, confirming the specificity of the method. The recombinant antigen's AChE phenotype was confirmed as Yta, as no reaction was detected with anti-Ytb-positive sera. The ELISA method correlated closely with the established serological grading system used routinely in blood transfusion laboratories.

Comparison of enzyme immunoassay and fluorescence polarization immunoassay as techniques for measuring anticonvulsant drugs on the same analytical instrument.

We have applied kits for enzyme immunoassay and fluorescence polarization immunoassay of anticonvulsant drugs to the same centrifugal analyser. There was a good correlation between the two techniques for the assay of phenytoin, carbamazepine, and phenobarbitone. Within- and between-batch reproducibility was also comparable but only the fluorescence polarization system allowed the use of stored calibration curves. The ability to use stored curves may be particularly advantageous to laboratories required to run a large number of stat assays or who are handling low workloads of particular analytes.

Effect of bilirubin on uricase-peroxidase coupled reactions. Implications for urate measurement in clinical samples and external quality assessment schemes.

Methods for the determination of serum urate which employ the uricase-peroxidase reaction may suffer interference from concentrations of bilirubin which can be found in relatively mild jaundice. Such concentrations of bilirubin are also frequently present in sera distributed as part of an external quality assessment scheme and, for this reason, laboratories should be particularly careful not to recalibrate their urate assays to attempt to achieve results closer to the consensus mean.

Evaluation of a commercial immunoenzymometric assay for alpha-fetoprotein.

A two-site immunoenzymometric assay (Abbott Diagnostics) for alpha-fetoprotein (AFP) in maternal serum and amniotic fluid has been evaluated for its suitability as a screening test for open neural tube defects. In a retrospective study based on 190 pregnancies of known outcome, performance of the kit in measuring both serum and amniotic fluid AFP correlated well with that of an in-house radioimmunoassay. Of 39 pregnancies associated with open neural tube defects, only four would not have been detected by the use of sequential measurement of serum and amniotic fluid AFP (also essentially in agreement with results obtained by the RIA). We conclude that this immunoassay could form the basis for a screening program for antenatal detection of open neural tube defects.

A prospective study of amniotic fluid cholinesterases: comparison of quantitative and qualitative methods for the detection of open neural tube defects.

The value of quantitative and qualitative methods of cholinesterase (ChE) analysis in the detection of open neural tube defect (NTD) has been assessed in a prospective survey of 1495 mid-trimester amniotic fluids. Using a quantitative method the mean ChE values were much lower in fluids from pregnancies of normal outcome but it was not possible to discriminate these fluids completely from those associated with NTD pregnancies, particularly when the specimens were contaminated with blood. Similarly, measurement of acetylcholinesterase (AChE) activity alone by three different methods also failed to eliminate the overlap between the two groups. In contrast, polyacrylamide gel electrophoresis revealed only a single band of ChE activity in 1408 out of 1410 fluids from pregnancies with a normal outcome whilst amniotic fluids from all 60 cases of open NTD, 6 out of 7 cases of exomphalos and 3 out of 4 cases of intra-uterine death gave the characteristic second faster-running AChE band. A qualitative gel method which requires the same amount of ChE activity to be loaded from each amniotic fluid is an effective method for pre-natal diagnosis of NTDs.

Amniotic fluid cholinesterase measurement as a rapid method for the exclusion of fetal neural tube defects.

Amniotic fluid total cholinesterase (ChE) and acetylcholinesterase (AChE) activities have been measured in 404 pregnancies without fetal malformation and 79 pregnancies associated with open neural-tube defects (NTDs). Neither measurement, either alone or in combination, gave complete separation of the two groups. However, measurement of ChE can be used to assess the probability that a woman is carrying a fetus with an open NTD. Since ChE can be measured within 15 minutes and shows at least 70% of women selected for amniocentesis by serum alpha-fetoprotein screening to have a less than 1 in 100 chance of carrying a fetus with an open NTD, we suggest that this test may be used in an amniocentesis clinic both to reassure women with normal pregnancies and to select probable abnormalities for immediate further investigation.

Purification, radioimmunoassay, and distribution of human brain 14-3-2 protein (nervous-system specific enolase) in human tissues.

Human 14-3-2 protein, a nervous-system specific enolase (EC 4.2.1.11) isoenzyme, has been purified from human brain and a sensitive radioimmunoassay has been developed for its detection. A systematic survey of human organs has shown that immunoreactive nervous-system specific enolase is present in all human organs but at levels less than 3% of those found in human brain, with especially low levels in liver, kidney and skeletal muscle, and with the highest levels in adrenal and large intestine. In all organs immunoreactive nervous-system specific enolase occurs in two forms representing the heterodimer and homodimer forms of the enzyme, and in all tissues except brain the heterodimer predominates. The presence of nervous-system specific enolase in other organs is unlikely to be explicable by innervation alone since significant quantities are found in red blood cell haemolysates. Tissues which contain amine precursor uptake and decarboxylation cells, for which the protein has been claimed to be a specific molecular marker, do not contain significantly higher levels of immunoreactive nervous-system specific enolase than other tissues. Both the heterodimer and homodimer forms of the enolase appear to be expressed at low levels in all tissues.