酶联免疫法检测饲料中己烯雌酚的探讨

己烯雌酚酶联免疫反应测试盒是用于饲料中己烯和其他相关的类固醇残留物如二氢己烯雌酚(hexe-strol)的定量检测。在家禽或鱼类中过量残留会对消费者造成危害,为此己烯雌酚已经被禁止在食品生产中使用。肉类食品中的过量残留,主要是由于被饲喂动物的饲料中含有高含量的己烯雌酚,动物缓慢蓄集所致,所以要想对食品中己烯雌酚得到有效控制,对饲料中的己烯雌酚进行监测至关重要。经实验,酶联免疫法具有操作简单、灵敏度高、时间短、检测成本低廉等优点,检测平均回收率≥98.3%,相关系数(r值)≥0.9992,离散系数(CV值)≤3.4%。     

Enzyme linked immunosorbent assay (ELISA) for the detection of antibodies to porcine cytomegalovirus.

The ELISA and indirect immunofluorescence test were compared on 56 porcine sera which were tested for antibodies to porcine cytomegalovirus. Viral antigens were prepared in cells of a pig fallopian tube line. The ELISA was found to be a sensitive reproducible and practical test to measure specific antibodies to this infection.     

Development of an enzyme linked immunosorbent assay (ELISA) for the detection of bovine serum antibody to bovine viral diarrhoea virus

An enzyme linked immunosorbent assay (ELISA) was developed to detect antibody to bovine viral diarrhoea virus (BVDV) in bovine serum. The ELISA results were compared with those of the serum neutralisation test (SNT) using serums from 6 experimentally infected calves bled at intervals from 0 to 154 days postinfection and 886 field samples. The optical density (OD) produced by a single dilution of test serum was compared with a standard curve and the result expressed in ELISA units. Despite wide variation between absolute ELISA and SNT results, an agreement of 97% was obtained when reciprocal SNT titres greater than or equal to 8 and ELISA units greater than or equal to 10 were taken as indicative of a specific reaction. The ELISA was shown to be an efficient method of measuring antibody in bovine serum samples and would assist in any large scale screening of cattle herds for BVDV antibody.     

Rapid detection of Chlamydia psittaci in vaginal swabs of aborted ewes and goats by enzyme linked immunosorbent assay (ELISA)

An enzyme linked immunosorbent assay (ELISA) for the detection of Chlamydia psittaci in vaginal swabs of aborted ewes and goats has been developed using microtiter plates coated with sheep anti-Chlamydia immunoglobulin G. This technique was compared to the direct isolation of the agent by plaque assay on McCoy cells. Among 89 specimens from animals in infected flocks, 58 were positive by both methods, seven were only positive by ELISA, and nine others were only positive by direct isolation (plaque assay). None of the 75 specimens from animals in healthy flocks gave a positive response in ELISA or the plaque assay. Unlike direct isolation in cell culture, the ELISA technique permitted the detection of Chlamydia even in the absence of special care in sampling and conservation of specimens.     

Seroepidemiology of rinderpest in bovids in Sri Lanka using the enzyme linked immunosorbent assay (ELISA) technique

Approximately 0.2% (n=4397) of the bovids (cattle and buffalo) in Sri Lanka were sampled, from June 1992 using a multi-stage sampling procedure. Serum antibodies for the rinderpest virus were detected using the competitive enzyme-linked immunosorbent assay. The age, the agroclimatic zone, the management system practiced in the farms, and the vaccination history of the sampled bovids were studied as potential risk factors for being seropositive.

The prevalence of rinderpest antibodies in non-vaccinated bovids was 3.5% (n=4101). The prevalence was higher in the dry zone (9%; where the outbreak emerged in 1987), compared to bovids in the other zones (1%). Seropositive bovids over three years of age were approximately at fourfold higher chances of being seropositive compared to those that were ≤3 years old. The higher prevalence in older animals is probably due to exposure to the virus during the 1987 epidemic. Bovids from the dry zone (annual rainfall 20 to 35 inches) were at higher odds of being seropositive even after controlling for the possible effects of age, agroclimatic zone, management system and vaccination. The fact that 62% of bovids from the dry zone in this study were reared under extensive management system (free grazing) which allow unrestricted contact between animals, may be the reason for the above finding. A relatively poor response to vaccination observed in vaccinated bovids (seroprevalence=12%; n=296) could be attributed to difficulties in maintaining the vaccine at recommended temperatures in the field. This is the first island-wide study on seroprevalence of rinderpest in Sri Lanka.     

Comparison of the enzyme-linked immunosorbent assay (ELISA) with three other methods for the detection of Trichinella spiralis infections in pigs

Four methods employed in the diagnosis of trichinellosis [trichinoscopy, digestion method, immunofluorescence techniques and Enzyme-Linked Immunosorbent Assay (ELISA)] were compared by laboratories in eight countries of the European Economic Community. Material from 24 pigs infected with 10 000, 5000, 500, 150 and 0 T. spiralis larvae was examined during a period from 17 days to 12 weeks post infection. ELISA was more sensitive than immunofluorescence during the onser of the infection in groups in infected with higher numberts of larvae (1500, 5000 and 10 000 larvae). In general, however, results of both ELISA and immunofluorescence were comparable with regard to reliability. In pigs infected with a lower number of T. spiralis larvae both serological assays were more sensitive than the direct methods (trichinoscopy and digestion method).It was concluded that not enough evidence was available to suggest ELISA as an alternative to the direct methods for slaughterhouse control. Both the ELISA and the immunofluorescence technique may prove to be applicable for epidemiological surveys.     

酶联免疫吸附测定法及在兽药饲料分析中的应用

酶联免疫吸附测定法(enzyme -linkedim munosorbentassay,简称ELISA)是2 0世纪70年代后迅速发展起来的免疫测定法,广泛用于人类、动物、植物以及微生物领域。近年来,随着兽药及兽药在饲料中的使用日益广泛,兽药种类及化学结构日趋复杂,以及检测的低剂量化、现场化和实验室样品的批量化,特别是人们对长期摄入低水平兽药可能导致的各种慢性及远期危害的关注,高选择性、高灵敏度、高分析效率、低成本、低环境污染特点的酶联免疫吸附测定法成为当今兽药饲料分析领域应用最广、发展最快的测定技术之一。1　酶联免疫吸附测定法的检测原理酶联免…     

Trichinella spiralis infections in pigs. Comparison of homologous passive cutaneous an aphylactic (PCA) reactions with the enzyme linked immunosorbent assay (ELISA)

Immunoglobulins were analysed in the sera of pigs inoculated once with different numbers of Trichenella spiralis. Analyses for IgE were performed by homologous passive cutaneous anaphylactic (PCA) reactions and for IgG (H + L) by enzyme linked immunosorbent assay (ELISA).In the higher inoculation dose range (5,000–10,000 larvae) IgE production paralleled that of IgG (H + L). In the lower range (150,500 and 1,500 larvae) IgE production preceded IgG production in most animals.The possible significance of this finding for the early diagnosis of T. spiralis infections is discussed.     

Development of an enzyme-linked immunosorbent assay (ELISA) for the detection of antibody to the parasitic dinoflagellate Amyloodinium ocellatum in Oreochromis aureus.

An enzyme-linked immunosorbent assay (ELISA) using antibody to affinity-purified Oreochromis aureus immunoglobulin and antigens from the parasitic dinoflagellate amyloodinium ocellatum was developed. The ELISA was then used to evaluate the immune response of the tilapine fish to immunization with the parasite. Fish immunized with antigens of the dinospore stage, either live or sonicated, produced a specific immune response that was detectable by this ELISA. Combinations of serial dilutions of A. ocellatum antigen and fish anti-A. ocellatum serum were examined to determine which dilutions provided optimal differentiation of seropositive from seronegative fish. Fresh and heat-inactivated serum from both seropositive and seronegative fish produced similar results.     

Rapid detection of viral-specific antibodies by enzyme-linked immunosorbent assay (ELISA)

The development of three separate rapid ELISAs for detecting antibodies in host serum to three different viruses is described. These include: 1. A direct antigen assay using enzyme labelled anti-canine Ig for detecting antibodies to canine parvovirus, 2. A competitive ELISA using a feline infectious peritonitis virus-specific monoclonal antibody labelled with enzyme, and 3. A competitive ELISA using an equine infectious anemia virus-specific monoclonal antibody and enzyme labelled antigen, p. 26. The utility and benefits of each of the three approaches is emphasized.     

An enzyme-linked immunosorbent assay (ELISA) for the detection of anti-chlamydial antibodies in pig sera

The evaluation of an enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies against chlamydiae in pig sera is described. The most widely used serological test is the complement fixation test (CFT). The CFT has a lack of sensitivity and specificity because of low antibody titers and unspecific reactions. Eight conventionally raised pigs were exposed to a pathogenic strain of Chlamydia suis, four controls were mock infected. The immune responses was monitored by CFT and indirect ELISA. There was no agreement between CFT and ELISA data. These results were confirmed by a study with 191 sera from nine pig farms. As shown by ELISA and PCR chlamydiae are widespread in swine.     

Assessment of an enzyme linked immunosorbent assay for the detection of cattle infected with Mycobacterium bovis

An indirect enzyme linked immunosorbent assay (ELISA) using an unpurified antigen was assessed for its accuracy in detecting Mycobacterium bovis infected cattle. The ELISA test recorded sensitivities of 88.7% and 63%, respectively, for infected cattle tuberculin tested positive and for infected cattle never tuberculin tested. Specificity was determined at 52.6% for cattle from confirmed free herds which had never been tuberculin tested. Significant differences in the mean ELISA values were recorded between the 3 groups. No evidence was found for long term effects of tuberculin testing upon the titre of antibodies detected by the ELISA in unaffected cattle. The indirect ELISA using the unpurified antigen of this assay was considered to be unsuitable as an alternative to tuberculin testing for the detection of M. bovis infected animals.     

Detection of serum antibodies in Eimeria tenella-infected chickens by enzyme linked immunosorbent assay (ELISA) with merozoite and oocyst antigens.

Soluble antigens prepared from sporulated oocytes and second generation merozoites of E. tenella were used for enzyme linked immunosorbent assay (ELISA) to investigate antibody in sera of two breeds of chickens, i.e. commercial broilers and SPF single comb white leghorn layers, which were experimentally infected with E. tenella. In broilers inoculated with oocysts at 15 days of age, ELISA values increased rapidly after day 19 post inoculation (PI) and reached the maximum lebel on days 29 and 32 PI against both merozoite and oocyst antigen. The values against merozoite antigen were significantly higher than those against oocyst antigen. In SPF layers infected at 15 days of age, the values increased gradually after 7 days PI. There were no significant differences between values against two antigens. Generally, the values in broilers tended to be higher than those in SPF layers, especially against merozoite antigen. In broilers inoculated with oocysts at 1 and 15 days of age, ELISA values increased rapidly and reached the maximum level on days 11 and 20 post second inoculation (PSI) against merozoite and oocyst antigens respectively and then the values against merozoite antigen decreased. The values against merozoite antigen were markedly higher than those against oocyst antigen. In SPF layers inoculated twice, the values reached the highest on day 11 PSI as in the case of broiler; however, after that day, the values against both antigens decreased. The sera reacted similarly against both antigens. The values against merozoite antigen were significantly higher in broilers than in SPF layers.(ABSTRACT TRUNCATED AT 250 WORDS)     

A dot enzyme linked immunosorbent assay (dot ELISA): Comparison with standard fluorescent antibody test (FAT) for the diagnosis of rabies in animals

A modified enzyme linked immunosorbent assay (Dot ELISA) is described for visual detection of rabies antigen in animals. The test materials were dotted onto the nitrocellulose paper and allowed to react with rabies antiserum. The bound antigen—anti-body were reacted with a peroxidase conjugated antirabbit immunoglobulin. Positive reactions were easily visualized as brown dots after enzyme degradation of the substrate. A total of 400 specimens from various geographical locations were tested with the dot ELISA technique, and also with the fluorescent antibody test (FAT), which was used as a reference method. The concordance between the two tests was 95.25%. The dot ELISA may have potential applications as a rapid, simple and economical field test in the diagnosis of rabies.     

Enhanced chemiluminescent enzyme immunoassay for the detection of trichinellosis antibodies in pigs.

A modified enhanced chemiluminescent enzyme assay (ECIA) was developed for mass screening of pigs for trichinellosis antibodies in abattoirs. Using Bionectics beads as solid support, the assay time could be reduced to 45 min. Optimal conditions for washing, blocking, incubation, concentration of serum, antigens and conjugates as well as timing of film exposure were determined. The sensitivity and specificity of the assay were found to be comparable to those of the triple antibody-IgG ELISA. The assay was tested in an abattoir and its efficacy was found to be satisfactory. However, the major disadvantage of the assay is the high cost of magnetic beads.     

An indirect enzyme linked immunosorbent assay for the detection of bovine antibodies to multiple Leptospira serovars

An indirect enzyme linked immunosorbent assay was developed for the detection of bovine antibodies to multiple pathogenic Leptospira serovars, including canicola, copenhageni (represents icterohaemorrhagiae), grippotyphosa, hardjobovis, pomona, and sejroe. The antigen utilized in this assay was a sonicated mixture of equal parts of killed whole cells of each of the 6 serovars named above. A mouse monoclonal antibody against bovine immunoglobulin (Ig)G₁ that was conjugated with horseradish peroxidase was used for detection of bound antibodies. This assay was evaluated with sera (n = 3107) that were microscopic agglutination test (MAT)-negative (at a 1:100 dilution) for each of the 6 serovars listed above and sera (n = 601) that were MAT-positive (at a 1:100 dilution) for 1, or any combination of the 6 listed serovars. In addition, sera from serial weekly bleedings of cows, which were individually experimentally infected with serovars hardjobovis, copenhageni, grippotyphosa, or canicola, were also tested in this assay.

At an optimal cut-off point determined by receiver operating characteristic (ROC) curve analysis, the relative sensitivity and specificity of the assay were 93.5% (95% confidence interval = 91.2% to 95.3%) and 94.7% (95% confidence interval = 93.9% to 95.5%), respectively. This assay was able to detect antibody in the sera of animals experimentally infected with serovar hardjobovis as early as 1 week postinoculation

     

Latest developments in the enzyme-linked immunosorbent assay (ELISA)

Enzyme-linked immunosorbent assays (ELISAs) for the serologic detection of both antigen and antibody in monitoring programs of commercial poultry flocks have begun to be recognized as an improvement over more conventional diagnostic procedures. The feasibility of employing double-antibody sandwich assays for the detection of virus without prior isolation of virus has been demonstrated and shows promise as the method of choice for the detection of lymphoid leukosis virus shedding. The versatility of indirect ELISA for the measurement of antibody induced by a wide variety of potential pathogens using a single basic overlapping ELISA system has also been demonstrated. It shows potential as a likely candidate to replace some of the more costly and time-consuming or less sensitive conventional serologic methods that do not overlap. Although some aspects of the two major types of immunoassays currently used in poultry health may need some modifications or improvements before delivery for routine use, it is likely that the use of computer-assisted ELISA will gain increased acceptance and use as the preferred way to efficiently and accurately monitor the health of poultry flocks on a broad scale.     

Development of a sandwich-dot enzyme linked immunosorbent assay for Streptococcus pneumoniae antigen detection in cerebrospinal fluid

A new sandwich dot-enzyme linked immunosorbent assay (sdot-ELISA) was developed using omniserum prepared against different strains of Streptococcus pneumoniae as capture antibody and also as second or revealing antibody after its conjugation with horseradish peroxidase (HRP) for detection of pneumococcal antigen in cerebrospinal fluid (CSF). A total of 103 CSF samples of different categories were screened with newly developed dot-ELISA and results were compared with commercially available latex agglutination (LA) kit. The newly developed sdot-ELISA was more sensitive than LA test and can be used as an alternative diagnostic tool in laboratory and in field conditions. An added advantage of this ELISA system was that it did not require antibodies produced in two different animal species.