Development of an ELISA assay for Clostridium perfringens phospholipase C (alpha toxin)

A new method for the assay of Clostridium perfringens alpha toxin (phospholipase C) is described using a sandwich ELISA. This assay has been shown to be quantitative, to have a high specificity for the toxin and is capable of detecting purified Clostridium perfringens phospholipase C at concentrations of as little as 0.005 units/ml in cooked meat culture medium.     

Development of an automated turbidimetric immunoassay for quantification of bovine serum immunoglobulin G

OBJECTIVE: To develop an automated turbidimetric immunoassay (TIA) for measurement of bovine IgG. SAMPLE POPULATION: 24 bovine serum samples. PROCEDURE: IgG concentration was determined by use of the TIA, and results were compared with those of the radial immunodiffusion (RID) method. Variables were determined, using commercially available reagents and a clinical biochemical analyzer. For the TIA, polyclonal goat anti-bovine IgG (Fc specific) serum, bovine IgG calibrator serum, and polyethylene glycol reaction buffer were used. Sample concentrations were determined by the instrument, using the linear regression method of least squares. The accuracy of this assay was validated by referencing to a purified bovine IgG standard and by recovery of control standards. Parallelism was documented by assay linearity and serial sample dilution linearity. Interference resulting from hemolyzed samples was examined. RESULTS: The TIA method correlated positively (r = 0.9957) and significantly (P < 0.05) with the RID method, yielding a regression equation with slope of 0.78708 and y-intercept of 1.02102. Bias attributable to hemolysis was not observed. CONCLUSIONS: The TIA method is automated, accurate, and precise for bovine serum IgG quantification. CLINICAL RELEVANCE: This assay provides sample results in approximately 10 minutes and may be used as an alternative to the manual RID method.     

An improved isolation technique for bovine Neospora species

The authors report on a case of a solitary liver abscess due to Listeria monocytogenes in a 53-year-old diabetic white male and review all published cases of solitary listerial abscesses of the liver. L. monocytogenes is a rare cause of solitary liver abscess which occurs in elderly patients with diabetes mellitus. The clinical signs are variable and often mimic malignancy, with epigastric pain, night sweats and weight loss. Prevalent features are poor control of glycemia, temperature up to 38.5 degrees C and elevated alkaline phosphatase. Optimal treatment includes percutaneous drainage of the hepatic abscess and antibiotic therapy with an aminopenicillin or trimethoprim/sulfamethoxazole. Outcome of the reviewed patients was favourable with no mortality and no relapse of the disease.     

Development of enzyme-linked immunosorbent assays using recombinant borrelial antigens for serodiagnosis of Borrelia burgdorferi infection

The basic tenet of continuous quality improvement is that there is always room for additional improvement in the clinical care provided to patients. The opportunities for this improvement come from the analysis of information collected during the ongoing monitoring of important elements of care. The provision of clinical psychiatric care is seen as a complex process that is dependent on the effective functioning of all of the health and mental health care organization. The concept of continuous quality improvement is the most recent stage in a long process of defining and redefining the basic goals and tenants of medical and psychiatric quality assurance. The determination of the actual improvement of psychiatric and mental health care due to quality assurance is a substantial and important technical problem. The determination of the value of this improvement in mental health care is an even greater ethical and social problem.     

Development of an ELISA to assess the potency of horse therapeutic polyvalent antibothropic antivenom

The objective of this study was the search for a suitable venom antigen to be used in an in vitro alternative immunoassay, to the standard antivenom neutralization assay using mice. Bothrops jararaca venom was fractionated in DEAE-Sephacel columns and the fractions were tested for a correlation between antibody capture enzyme linked immunosorbent assay (ELISA) absorbance values and the 'in vivo' antivenom potency. Individual antivenoms from 14 horses and 15 separate FUNED polyspecific Bothrops ampouled antivenoms (final product) were used. Fractions showing the higher correlations were further chromatographed in a Sephadex G-75 column and again tested for the correlation. Two fractions with haemorrhagic activity displayed a correlation of r = 0.77 and r = 0.8 against the individual horse antivenom sera and of r = 0.79 and r = 0.8 for the ampouled antivenom. For all results p < 0.001. Two other fractions with phospholipase A2 activity showed a correlation of r = 0.66 (p < 0.01) and r = 0.56 (p < 0.03) against the individual horse antivenom sera. Electrophoresis results show a similar composition for both antigens with haemorrhagic activity. Results indicate that the fractions purified would be suitable for the desired objective of this study.     

Development of an ELISA for pantothenic acid (vitamin B5) for application in the nutrition and biological fields

Immunological assays appear to be the only alternative to the microbiological method for analysis of pantothenic acid in foods and blood. In order to evaluate the influence of the linker on the immunogenicity of the hapten, we have tried to raise antisera against pantothenic acid in rabbits using different conjugates. The hapten was coupled to a carrier protein (BSA or thyroglobulin) using adipoyl dichloride (adipoyl conjugate) or bromoacetyl bromide (acetyl conjugate). Only the acetyl conjugate has induced the production of a specific antibody. With this antibody, an assay on microplate using the ELISA inhibition technique was developed to measure pantothenic acid. The use of pantothenic acid coupled to thyroglobulin with adipoyl dichloride as the capture antigen has improved the sensitivity of the ELISA. This assay was applied to food products and blood.     

延边黄牛气肿疽间接ELISA诊断方法的建立

[目的]检测延边黄牛气肿疽病原体,为牛气肿疽的检测提供科学方法和依据.[方法]采用气肿疽梭菌裂解菌体为抗原,建立检测该病原菌的酶联免疫吸附检测方法,确定其最适工作条件.[结果]气肿疽梭菌菌体裂解抗原最适包被浓度为50 μg/ml;待检血清的最适稀释度为1∶ 200,酶标二抗的最适工作浓度为1∶ 2 000;对随机采集于延边地区的30份牛血清样品进行间接ELISA检测,阳性率为20%.[结论]建立的间接ELISA方法特异性和重复性好.     

Survival of Mycobacterium paratuberculosis and preservation of immunoglobulin G in bovine colostrum under experimental conditions simulating pasteurization

OBJECTIVE: To determine whether Mycobacterium paratuberculosis could survive in colostrum after pasteurization. Additionally, this study investigated the effect pasteurization had on IgG concentration in colostrum. ANIMALS: Colostrum samples were collected from cattle (beef and dairy) owned by the state of Ohio. PROCEDURE: Colostrum was divided into aliquots and inoculated with variable concentrations of M paratuberculosis (ATCC No. 19698: 10(4), 10(3), and 10(2) colony-forming units/ml). Half the samples at each concentration were subjected to pasteurization temperatures (63 C) for 30 minutes and the remainder were kept at approximately 20 to 23 C. All samples were incubated (Herrold's egg yolk medium with and without mycobactin J) and observed for growth during the next 16 weeks. Additionally, the IgG concentration of colostrum was determined by radioimmunoassay before and after pasteurization. Samples that coagulated at pasteurization temperatures were mechanically resuspended before measurement of IgG concentration. RESULTS: Growth of M paratuberculosis was retarded but not eliminated by pasteurization. Growth was observed in all unpasteurized samples incubated on Herrold's egg yolk medium with mycobactin J but in only 2 of 18 pasteurized samples similarly cultured. Growth from pasteurized samples appeared 5 to 9 weeks after growth was observed from nonpasteurized samples. Mean colostral IgG concentration was 44.4 g/L in nonpasteurized samples and 37.2 g/L in pasteurized samples, a decrease of 12.3%. High-quality colostrum (> 48 g of IgG/L) had a significantly greater loss of IgG concentration than did colostrum of lesser quality (P = 0.002). CONCLUSIONS: Pasteurization lessened, but did not eliminate, growth of M paratuberculosis from experimentally inoculated colostrum samples. Pasteurization resulted in a significant decrease in colostral IgG concentration but not to an unmanageable level that would preclude the colostrum's use for passive transfer of immunity. CLINICAL RELEVANCE: Colostrum is macrophage rich and may serve as a source of M paratuberculosis infection to calves. Pasteurization of colostrum may lessen the risk of infection, but will not totally eliminate M paratuberculosis.     

Transmission of paratuberculosis

Antenatally, as more women are being treated in the home setting rather than the hospital setting, diagnostic modalities such as ultrasonographic imaging need to be available for use by the home care provider. In the home setting, the care provider and treatment modalities are brought to the patient, whereas patients managed in the hospital setting must attend the provider site to receive the same services. Without the availability of diagnostic ultrasonography in the home setting, pregnant women who are treated in the home rather than the hospital are required to incur an added financial and physical burden associated with attendance at care. Financially, nonreimbursed out-of-pocket costs incurred by the patient may include costs of transportation, costs of child care, and loss of wages. Physically, patients are prevented from maintaining recommended bed rest and dietary treatment regimens. Ultrasonographic evaluations can be performed safely in the home setting by nurses who have received added educational preparation and ultrasonographic imaging experience, thereby providing the essential care often necessary for women who experience pregnancies complicated by high-risk conditions.     

Development of a sensitive ELISA for human leptin, using monoclonal antibodies

A new, sensitive ELISA for human leptin in plasma and cerebrospinal fluid (CSF) was developed, using monoclonal antibodies. The lower limit of detection of this ELISA was 0.78 pg/assay. Both intra- and interassay imprecision values were <7%. The dilution curves of plasma and CSF showed good linearity, and the recovery was 83.2-95.6%. There was good correlation between plasma leptin concentrations by the ELISA and a commercially available RIA (r = 0.99). Our ELISA is advantageous because it does not require radioisotopes, it produces results in hours rather than days, and more importantly, it improves on the detection limit and plasma interference of the RIA kit. The new ELISA enables measurement of low concentrations of leptin, as are seen in CSF and in plasma of patients with anorexia nervosa.     

A study for serodiagnosis of verotoxin-producing Escherichia coli (enterohemorrhagic E. coli) O157 by the bacterial agglutination technique

We evaluated the usefulness of bacterial agglutination antibodies for serodiagnosis of verotoxin-producing Escherichia coli (enterohemorrhagic E. coli) O157 infections. We examined 50 serum samples from 50 control children (whiout diarrhea 31, with diarrhea 19), 24 samples from 8 diarrhea cases due to O157:H7, 37 samples from 14 cases of hemolytic uremic syndrome (HUS) for antibodies to heat-killed E. coli E32511 (O157:H.-) strain using the bacterial agglutination technique. Of the control sera all but one (x80) showed 20 > or = in the antibody. All the diarrhea patients due to O157:H7 showed a significant rise (x160-x5120) of the titers in the sera at 5-7 days on illness, after that the titers fell rapidly. Significant antibody rise (x160-x5120) was detected in twelve out of 14 HUS patients at the early stage of the illness which fell in the convalescent phase. The assay appeared to be a useful serodiagnostic technique because of its easiness and simplicity as well as because of its high sensitivity and specificity.     

Detection of anti-Histoplasma capsulatum antibodies by the ELISA technique. Preliminary study

An indirect micro-ELISA system is presented for diagnosing histoplasmosis. The diagnostic criteria are defined by using sera from 12 patients who are histoplasmosis carriers. For this group, the optical density values were superior to 1,000; use was made of 43 sera from blood bank donors and 9 sera from children without a history of exposure. The optical density values in these cases were inferior to 0,200. The significant difference found led to the diagnostic criterion for confirming 3 histoplasmosis carriers who showed clinical manifestations but had been negative to double immunodiffusion. Thus, the usefulness of the proposed micro-ELISA system for early diagnosis was proved.     

Development of sera reference panels for quality control of ELISA diagnostic kits in Russia

The clinico-immunological examination of 57 patients with chronic bacterial prostatitis was carried out. The clinical analysis made it possible to divide the patients into 3 groups characterized by the presence of chronic inflammatory diseases of other organs which had appeared before (group 1) or after (group 2) the manifestation of the symptoms of prostatitis, aw well as by the absence of concomitant inflammatory diseases (group 3). At the same time these patients were found to have changes in their immune status, most pronounced in patients of groups 1 and 2. The clinico-immunological analysis of the patients with chronic bacterial prostatitis revealed the fact that chronic bacterial prostatitis was a chronic inflammatory process linked with changes in the immune system; these changes had the signs of secondary immunodeficiency and required immunocorrective therapy.     

Peptide-based OspC enzyme-linked immunosorbent assay for serodiagnosis of Lyme borreliosis

Sera from 210 patients with Lyme borreliosis (LB) were studied by an enzyme-linked immunosorbent assay (ELISA) based on a synthetic peptide (pepC10) comprising the C-terminal 10-amino-acid residues of OspC of Borrelia burgdorferi. We found that 36.3 and 45.0% of the serum samples from patients with erythema migrans (EM) and neuroborreliosis (NB), respectively, displayed immunoglobulin M (IgM) anti-pepC10 reactivities, while these samples rarely (相似文献   

宝钢数字化自动煤岩分析技术

多学科合作开发的数字化自动煤岩分析技术,采用高精度和可改变曝光时间数码摄像头,扩展了灰度级, 提高了反射率测定精度。利用所获高分辩率显微图像,采用图像识别技术,实现煤岩组分的自动识别分类。结合自动扫描、自动聚焦,实现了煤岩特性的自动快速分析。克服了显微光度计速度慢和图像分析仪精度低的不足,继承了显微光度计精度高和图像分析仪的测定速度快的优点,使得测定结果准确、可靠。实现了煤岩特性测定的数字化和自动化,速度快、劳动强度低,操作人员不要经过专业培训。     

Assessment of intact Sarcocystis cystozoites as an ELISA antigen

Hereditary hemorrhagic telangiectasia (HHT), or Rendu-Osler-Weber disease, is an autosomal dominant disorder characterized by a triad of mucocutaneous and visceral telangiectasia, recurrent epistaxis and familial history. We reported a rare case of HHT associated with pulmonary and cerebral arteriovenous fistulae (AVF) and multiple cerebral arteriovenous malformations (AVM). The roles of multimodality therapies including artificial embolization, feeder clipping and stereotactic radiosurgery for these multiple cerebrovascular dysplasia in HHT were discussed. In particular the usefulness of radiosurgery to obliterate AVM was emphasized. It is especially useful for multiple AVM's associated with HHT. A 7-year-old boy had presented himself at another hospital 2 years previously with cyanosis of the lips and fingers on exertion. He was diagnosed as having pulmonary AVG and underwent surgery. His mother had suffered from epistaxis in her adolescence, and was then highly suspected as having HHT. She underwent surgical removal of a left fronto-parietal AVM at the age of 16 years. The family history then prompted the patient to have a brain CT done, which eventually demonstrated an abnormal enhancing mass at the left frontal region. He was transferred to our service for further evaluation. Left carotid angiograms demonstrated an AVF supplied by a dilated anterior internal frontal artery of the anterior cerebral artery (ACA), draining directly into the vein of the corpus callosum with a large aneurysmal dilatation, and then draining further into the straight sinus via the vein of Galen. In addition, right carotid angiograms revealed three small AVM's fed by the median artery of the corpus callosum, and the middle internal frontal and paracentral arteries of the right ACA, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

Development of PCR tests for the detection of bovine herpesvirus-1, bovine respiratory syncytial viruses and pestiviruses

The development of PCR assays for detection of BHV-1, BRSV, BVDV and another pestiviruses is summarized. A polymerase chain reaction assay based on primers selected from the viral gI glycoprotein gene detected 3 fg pure BHV-1 DNA, 0.1-1.0 TCID50 or a single infected cell. No amplification was observed with DNA from BHV-2, BHV-3, BHV-4, OHV-1 or OHV-2. However, a fragment of the correct size (468 bp) was amplified using DNA from herpesviruses isolated from reindeer, red deer and goat. The PCR assay was able to detect virus in nasal swabs 1-14 days after experimental infection of cattle and there was a good correlation when PCR was compared to virus isolation for the detection of BHV-1 in clinical field samples. Detection of BHV-1 in fetal bovine serum and semen samples was also successful. PCR detecting a broad range of BVDV, BDV and HCV was developed. Of six sets of primers selected from different parts of the pestivirus genome the best results were provided by a pair 324/326 from the highly conserved 5'-non-coding region which gave an amplification with all 129 isolates tested. This panel consisted of 79 isolates from cattle, 33 from pigs and 17 from sheep. Differentiation between viruses was achieved by cleavage of the PCR-amplified products (288 bp) with the restriction endonucleases AvaI and BglI. The BVDV products were cleaved by AvaI, HCV by BglI and AvaI. Both enzymes, AvaI and BglI, did not cut the BDV products. A nested polymerase chain reaction assay was developed for the detection of bovine respiratory syncytial virus (BRSV). Primers were selected from the gene encoding the F fusion protein. The sensitivity of PCR assay was 0.1 TCID50. No cross reaction was observed with nine heterologous respiratory viruses. PCR products of bovine and human RSV strains were discriminated using endonuclease ScaI, which specifically cleaved products of BRSV. PCR assay detected BRSV in nasal swabs collected from cattle in the acute stage of respiratory disease. In vitro amplification detected 31 positive samples of 35 while immunofluorescence only 23 samples.