Efficacy of Several Serological Tests and Antigens for Diagnosis of Bovine Brucellosis in the Presence of False-Positive Serological Results Due to Yersinia enterocolitica O:9

Yersinia enterocolitica O:9 bears a smooth lipopolysaccharide (S-LPS) of Brucella sp. O-chain A + C/Y epitopic structure and is a cause of false-positive serological reactions (FPSR) in standard tests for cattle brucellosis. Brucella S-LPS, cross-reacting S-LPSs representing several O-chain epitope combinations, Brucella core lipid A epitopes (rough LPS), Brucella abortus S-LPS-derived polysaccharide, native hapten polysaccharide, rough LPS group 3 outer membrane protein complexes, recombinant BP26, and cytosolic proteins were tested in enzyme-linked immunosorbent assays (ELISA) and precipitation tests to detect cattle brucellosis (sensitivity) and to differentiate it from FPSR (specificity). No single serological test and antigen combination showed 100% sensitivity and specificity simultaneously. Immunoprecipitation tests with native hapten polysaccharide, counterimmunoelectrophoresis with cytosolic proteins, and a chaotropic ELISA with Brucella S-LPS were 100% specific but less sensitive than the Rose Bengal test, complement fixation, and indirect ELISA with Brucella S-LPSs and native hapten or S-LPS-derived polysaccharides. A competitive ELISA with Brucella S-LPS and M84 C/Y-specific monoclonal antibody was not 100% specific and was less sensitive than other tests. ELISA with Brucella suis bv. 2 S-LPS (deficient in C epitopes), Escherichia hermannii S-LPSs [lacking the contiguous α-(1-2)-linked perosamine residues characteristic of Y. enterocolitica S-LPS], BP26 recombinant protein, and Brucella cytosolic fractions did not provide adequate sensitivity/specificity ratios. Although no serological test and antigen combination fully resolved the diagnosis of bovine brucellosis in the presence of FPSR, some are simple and practical alternatives to the brucellin skin test currently recommended for differential diagnosis.     

Novel Recombinant-Antigen Enzyme Immunoassay for Serological Diagnosis of Syphilis

Enzyme immunoassay (EIA) is an ideal method for screening large numbers of patients for syphilis. We evaluated a novel immune-capture EIA (ICE Syphilis; Murex Diagnostics) that uses three recombinant Treponema pallidum antigens (TpN15, TpN17, and TpN47) and compared the results with those obtained by the native T. pallidum antigen EIA (Captia SelectSyph-G; Centocor) that we currently use for the serodiagnosis of syphilis. Specificity was evaluated by screening 1,184 unselected serum specimens in parallel by the ICE Syphilis and SelectSyph-G assays, while sensitivity was tested with a panel of 101 serum specimens containing antitreponemal antibodies (treated and untreated) from patients with various stages of infection. The specificity of the ICE Syphilis EIA (99.8%) on screening was significantly higher (P < 0.02) than that of the SelectSyph-G EIA (99.2%). The sensitivity of the ICE Syphilis EIA was significantly higher (P < 0.01) than that of the SelectSyph-G EIA on both initial (99 versus 91.4%) and repeat (100 versus 92.4%) testing. The ICE Syphilis EIA was also significantly more sensitive (P < 0.01) than the fluorescent treponemal antibody-abs (92.4%) but not the T. pallidum hemagglutination assay (97.1%). Sera containing antitreponemal antibodies gave a much higher antibody index (absorbance of test serum/kit cutoff) by the ICE Syphilis EIA than by the SelectSyph-G EIA. This combined with the overall high sensitivity makes the ICE Syphilis EIA an ideal test for excluding or detecting treponemal infection in human immunodeficiency virus (HIV)-infected patients. The ICE Syphilis EIA was positive with sera from all 15 HIV-infected patients in the study, whereas sera from 3 HIV-infected patients were negative by the SelectSyph-G EIA. We conclude that the high sensitivity and specificity of the ICE Syphilis EIA and its suitability for automation make it an ideal screening test.     

Validation of Immune-Complex Enzyme Immunoassays for Diagnosis of Pneumococcal Pneumonia among Adults in Kenya

The efficacy of pneumococcal vaccines in protecting against pneumococcal pneumonia can feasibly be measured only with a diagnostic technique that has a high specificity (0.98 to 1.00) and a sensitivity greatly exceeding that of blood cultures (>0.2 to 0.3). In this context immune-complex enzyme immunoassays (EIAs) offer a novel, convenient diagnostic method, and we have investigated three such assays with appropriate study populations in Kenya. Sera from 129 Kenyan adults with pneumococcal pneumonia and 97 ill controls from the same clinics, but without pneumococcal disease syndromes, were assayed with immune-complex EIAs for pneumolysin, C-polysaccharide, and mixed capsular polysaccharides (Pneumovax II). At an optical density (OD) threshold yielding a specificity of 0.95, the sensitivities (95% confidence intervals) of the assays were 0.22 (0.15 to 0.30), 0.26 (0.19 to 0.34), and 0.22 (0.15 to 0.29), respectively. For pneumolysin immune complexes, human immunodeficiency virus (HIV)-positive patients had a higher mean OD than HIV-negative patients (639 versus 321; P < 0.0001), but stratification by HIV infection status did not alter the performance of this test. Combining the results of all three EIAs did not enhance the diagnostic performances of the individual assays. In Kenyan adults the sensitivities of the immune-complex EIAs could exceed that of blood cultures only at levels of specificity that were insufficient for the performance of vaccine efficacy studies.     

Serological,Bacteriological, and Molecular Diagnosis of Brucellosis in Domestic Animals in Croatia

Aim

To present the surveillance data on Brucella melitensis, B. suis, and B. ovis infection in cattle, sheep, goats, and swine in Croatia obtained in 2008 by serological, bacteriological, and molecular methods for diagnostics of brucellosis in domestic animals.

Methods

We serologically tested 42 785 cattle serums, 22 686 sheep and goat serums, and 28 520 swine serums using the Rose Bengal test, complement fixation test, and various immunosorbent assays. We also tested 10 173 ram blood samples for B. ovis infection using the complement fixation test. Bacteriological examination was conducted on 214 samples collected from 34 serologically positive animals. Different molecular methods were employed in the identification and typing of 20 isolates from the samples.

Results

B. melitensis biovar (bv.) 3 was confirmed with different identification methods in 2 flocks in 2 Croatian counties and B. suis bv. 2 in 3 herds in 3 counties. B. melitensis in cows was confirmed for the first time in Croatia. Infection with B. ovis was serologically confirmed in 202 rams in 12 counties.

Conclusions

In 2008, the size of the brucellosis-affected area in Croatia and the efficiency of detection and prevention of brucellosis in sheep, goats, and swine were satisfactory. Infection with B. melitensis in cattle was confirmed for the first time and possible links for infection in humans were detected. More efficient measures for suppression and control of ovine epididymitis are required and a new strategy may be necessary for complete eradication of this disease.Brucellosis is a chronic infectious disease caused by bacteria of the genus Brucella that affects animals and humans. Each species of Brucella has their preferred host: B. abortus infects cattle, B. metitensis sheep and goats, B. suis swine, B. canis dogs, and B. ovis sheep, although they can also infect other animals (1). Brucellosis in sheep and goats is endemic in the Mediterranean region but is spread throughout Asia, Africa, and Central and South America (2,3). Along with tuberculosis and rabies, brucellosis is the most important bacterial zoonosis and remains an important public health and economic concern.With the exception of B. ovis and B. neotomae, all Brucella species can cause infections in humans. New Brucella species pathogenic for humans – B. ceti and B. pinnipedialis – have recently been discovered in marine mammals (4). Infection is transmitted to humans though direct contact with the infected animals or by consuming infected milk or fresh cheese (1).In Croatia, brucellosis in domestic animals is controlled in accordance with the annual order issued by the Ministry of Agriculture. Serological blood examination of all male breeding animals is mandatory twice per year, and all cases of abortion must be reported and tested for brucellosis. On large cattle and pig farms, 20% of breeding animals must be tested annually. Castration of seropositive rams without the obligation of bacteriological testing is required as an eradication measure for B. ovis infection.Bovine brucellosis (B. abortus) was eradicated in Croatia in 1964, while brucellosis in sheep and goats has occurred sporadically in the recent years, limited to 1-2 sheep flocks per year. All of the occurrences have resulted from epizooty originating in the neighboring country of Bosnia and Herzegovina (BH) (5,6). Swine brucellosis has been detected in swine and wild boars during regular controls (7,8) and B. suis isolates were determined as biovars (bv.) 1, 2, or 3 (7-11).B. ovis in rams and sheep causes either clinical or subclinical disease and is not pathogenic for humans (12). According to simulation models, B. ovis infection causes significant economic losses in flocks with no control measures, but there is no exact confirmation of the extent of such losses (13,14). Eradication is possible, but requires considerable resources.The aim of this study was to determine the effectiveness of the existing programs for diagnosis and control of brucellosis in domestic animals in order to prevent transmission of disease to humans and to reduce economic losses in animal production. This article describes the spread of brucellosis caused by B. melitensis, B. suis, and B. ovis in cattle, sheep, goats, and swine in the Republic of Croatia in 2008, as determined using different diagnostic methods.     

Enzyme Immunoassays for Measurement of Cytomegalovirus Immunoglobulin M Antibody

The diagnosis of congenital cytomegalovirus (CMV) infection is often accomplished by the detection of circulating antibody directed against CMV. We devised a method for measuring CMV-specific immunoglobulin M (IgM) based on the isolation of IgM antibody by reaction with a solid phase coated with antihuman IgM. The determination of IgM antibody specific for CMV was accomplished by the subsequent addition of CMV or control antigen and enzyme-labeled CMV antibody (solid phase-IgM method). We compared the sensitivity and specificity of this method with those of a conventional form of solid-phase enzyme immunoassay in which CMV antigen is bound to the solid phase (solid phase-antigen method). Both assay systems were capable of detecting CMV-specific IgM antibody in the sera of 10 babies with documented CMV infection and in those of the mothers of 4 of these babies. The solid phase-IgM method yielded negative results in all 66 sera available from babies who did not have congenital CMV infection. On the other hand, the solid phase-antigen system yielded false-positive results in 12 (18%) of these sera. In addition, the solid phase-antigen system yielded false-positive results in 8 of 12 sera obtained from patients with demonstrable rheumatoid factor. However, the solid phase-IgM system yielded negative results for the rheumatoid sera, provided that appropriate control reactions were performed. The solid phase-IgM system is thus a specific and sensitive method for the determination of CMV IgM antibody.     

Hepatitis E virus infection in Latin America: A review

Data reported during recent years reveal the complex picture of the epidemiology of hepatitis E virus (HEV) infection in Latin America. Whereas in countries like Argentina and Brazil is almost identical to the characteristic of most countries from North America and Europe, HEV in the Caribbean and Mexico involves the water‐borne, non‐zoonotic viral genotypes responsible for epidemics in Asia and Africa. Nevertheless, Latin America has been considered a highly endemic region for hepatitis E in the scientific literature, a generalization that ignores the above complexity. In addition, reports from isolated Amerindian communities, which display well known, important and very specific epidemiological features for hepatitis B and D virus infections are neither taken into account when considering the epidemiology of hepatitis E in the region. This review updates compilation of the available information for the HEV infection, both among humans and other mammals, in Latin America, discusses the strengths and the weaknesses of our current knowledge, and identifies future areas of research. J. Med. Virol. 85: 1037–1045, 2013. © 2013 Wiley Periodicals, Inc.     

Evaluation of Two Enzyme Immunoassays for Detection of Human Rotaviruses in Fecal Specimens

The two assays evaluated in this study (the Ridascreen rotavirus and the Pathfinder rotavirus) exhibited comparable sensitivities (100%) but highly divergent positive predictive values (93.74 and 57.7%, respectively) when compared on 393 specimens. This difference should be considered when using these tests on collectives with an unknown or low prevalence.     

mantisGRID: A Grid Platform for DICOM Medical Images Management in Colombia and Latin America

This paper presents the mantisGRID project, an interinstitutional initiative from Colombian medical and academic centers aiming to provide medical grid services for Colombia and Latin America. The mantisGRID is a GRID platform, based on open source grid infrastructure that provides the necessary services to access and exchange medical images and associated information following digital imaging and communications in medicine (DICOM) and health level 7 standards. The paper focuses first on the data abstraction architecture, which is achieved via Open Grid Services Architecture Data Access and Integration (OGSA-DAI) services and supported by the Globus Toolkit. The grid currently uses a 30-Mb bandwidth of the Colombian High Technology Academic Network, RENATA, connected to Internet 2. It also includes a discussion on the relational database created to handle the DICOM objects that were represented using Extensible Markup Language Schema documents, as well as other features implemented such as data security, user authentication, and patient confidentiality. Grid performance was tested using the three current operative nodes and the results demonstrated comparable query times between the mantisGRID (OGSA-DAI) and Distributed mySQL databases, especially for a large number of records.     

Performance of Two Commercially Available Automated Immunoassays for the Determination of Epstein-Barr Virus Serological Status

This study evaluated the performance of two automated Vidas (V) and Liaison (L) immunoassays for Epstein-Barr virus (EBV) serology. The detection of the viral capsid antigen (VCA) IgM, the VCA/early antigen (VCA/EA) IgG, and the Epstein-Barr nuclear antigen (EBNA) IgG was assessed on 526 sera collected for routine EBV testing in immunocompetent subjects. The determination of expected EBV status (186 EBV primary infections, 183 past EBV infections, and 157 EBV-seronegative individuals) was based on results of routine laboratory enzyme immunoassays (EIAs) together with clinical data. The sensitivity and specificity of each individual marker were determined in comparison to the expected EBV status. The agreement between the V and L profiles and the expected EBV status was established through the interpretation of combinations of the different EBV markers. Statistically significant differences between the two tests were found for the specificity of the VCA IgM marker (96.2% for V versus 93.2% for L), the sensitivity of the VCA/EA IgG marker (89% for V versus 94% for L), and the specificity of the EBNA IgG marker (96.5% for V versus 74.2% for L). The results determined for the two assays with respect to overall agreement with the established expected EBV status were not significantly different (89.7% for V versus 88.2% for L), with discrepancies mainly observed in sera referenced as primary infections. These findings demonstrated the similar performances of the Vidas and the Liaison assays for the establishment of an EBV serological status using the VCA, EA, and EBNA markers.     

Application of a Polymerase Chain Reaction Enzyme Immunoassay in Peripheral Whole Blood and Serum Specimens for Diagnosis of Acute Human Brucellosis

A simple polymerase chain reaction-enzyme immunoassay (PCR-EIA) was employed for the rapid laboratory diagnosis of human brucellosis directly from peripheral blood. Whole blood and serum specimens were collected from 243 patients with acute brucellosis as determined by blood culture, serological tests, and the patients clinical characteristics and from a control group of 50 healthy individuals. Diagnosis of brucellosis was established in 179 cases by isolation of Brucella spp. in blood culture and in 64 cases by clinical signs and serological investigation. Following the amplification of a 223-bp sequence of a gene that codes for the synthesis of an immunogenic membrane protein specific for the Brucella genus, the amplified product was detected in a microtiter plate by hybridization. Two hundred forty-one of the 243 patients tested had detectable Brucella DNA in either whole blood or serum specimens: 149 (61.3%) patients were positive in both whole blood and serum specimens, 43 (17.7%) were positive in serum specimens only, and 49 (20.2%) were positive in whole blood specimens only. The diagnostic specificity of the PCR-EIA assay for both specimen categories was 100%, while the sensitivity was 81.5% for whole blood specimens, 79% for serum specimens, and 99.2% for whole blood and serum specimens combined. The results suggest that the detection of Brucella DNA in whole blood and serum specimens by PCR-EIA assay is a sensitive and specific method that could assist the rapid and accurate diagnosis of acute human brucellosis.     

Diagnosis of Brucellosis in Livestock and Wildlife

Aim

To describe and discuss the merits of various direct and indirect methods applied in vitro (mainly on blood or milk) or in vivo (allergic test) for the diagnosis of brucellosis in animals.

Methods

The recent literature on brucellosis diagnostic tests was reviewed. These diagnostic tests are applied with different goals, such as national screening, confirmatory diagnosis, certification, and international trade. The validation of such diagnostic tests is still an issue, particularly in wildlife. The choice of the testing strategy depends on the prevailing brucellosis epidemiological situation and the goal of testing.

Results

Measuring the kinetics of antibody production after Brucella spp. infection is essential for analyzing serological results correctly and may help to predict abortion. Indirect ELISAs help to discriminate 1) between false positive serological reactions and true brucellosis and 2) between vaccination and infection. Biotyping of Brucella spp. provides valuable epidemiological information that allows tracing an infection back to the sources in instances where several biotypes of a given Brucella species are circulating. Polymerase chain reaction and new molecular methods are likely to be used as routine typing and fingerprinting methods in the coming years.

Conclusion

The diagnosis of brucellosis in livestock and wildlife is complex and serological results need to be carefully analyzed. The B. abortus S19 and B. melitensis Rev. 1 vaccines are the cornerstones of control programs in cattle and small ruminants, respectively. There is no vaccine available for pigs or for wildlife. In the absence of a human brucellosis vaccine, prevention of human brucellosis depends on the control of the disease in animals.Brucellae are Gram-negative, facultative intracellular bacteria that can infect many species of animals and man. Ten species are recognized within the genus Brucella. There are 6 “classical” species: Brucella abortus, Brucella melitensis, Brucella suis, Brucella ovis, Brucella canis, and Brucella neotomae (1,2). This classification is based mainly on differences in pathogenicity and host preference (3). Distinction between species and between biovars of a given species is currently performed using differential tests based on phenotypic characterization of lipopolysaccharide (LPS) antigens, phage typing, dye sensitivity, requirement for CO₂, H₂S production, and metabolic properties (1,2).The main pathogenic species worldwide are B. abortus, responsible for bovine brucellosis; B. melitensis, the main etiologic agent of ovine and caprine brucellosis; and B. suis, responsible for swine brucellosis. These 3 Brucella species cause abortion (“abortion storm” in naive heifers), and when brucellosis is detected in a herd, flock, region, or country, international veterinary regulations impose restrictions on animal movements and trade, which result in huge economic losses. These are the reasons why programs to control or eradicate brucellosis in cattle, small ruminants, and pigs have been implemented worldwide (4).B. ovis and B. canis are responsible for ram epididymitis and canine brucellosis, respectively. In the case of B. neotomae, only strains isolated from desert wood rat (Neotoma lepida) in North America have been reported. Recently 4 new Brucella species have been described: Brucella pinnipedialis and Brucella ceti, isolated predominantly from seals and cetaceans, respectively (5); Brucella microti, isolated from common voles (Microtus arvalis) (6), soil (7), and foxes (Vulpes vulpes) (8); and Brucella inopinata, isolated from a breast implant (9).There is a general host restriction pattern among the different Brucella species, meaning that different Brucella species infect different preferred hosts. Even within the B. suis species, different biovars preferentially infect different animal host species (1-3). Indeed, B. suis biovars 1 and 3 infect suidae, biovar 2 infects suidae and hare (Lepus europeanus), biovar 4 infects reindeer (Rangifer tarandus tarandus) and caribou (Rangifer tarandus granti), and biovar 5 has been isolated from rodents in Russia.All Brucella species may also infect wildlife species. Classical Brucella species have been isolated from a great variety of wildlife species such as bison, elk, feral swine, wild boar, fox, hare, African buffalo, reindeer, and caribou (10). In order to implement appropriate control measures to address wildlife brucellosis, it is very important to distinguish between a spill-over of infection contracted from domestic animals and a sustainable infection (10). In the latter case, the concern of the livestock industry is to prevent the re-introduction of the infection in livestock (spill-back), particularly in regions or states that are “officially brucellosis-free.” If the status of “officially brucellosis-free” is lost, domestic animals must be tested prior to being traded, which imposes huge costs. This is exemplified by recent episodes of cattle being infected with B. abortus transmitted by elk in the Greater Yellowstone Area in the USA (11) and of outdoor reared pigs infected with B. suis biovar 2 transmitted by wild boar in France (12).Brucellosis is an established zoonosis: infections have been attributed to at least 5 of the 6 classical Brucella species in terrestrial mammals. Studies from around the world indicate that elimination of the animal brucellosis reservoir has resulted in a substantial decline in the incidence of human disease (13). Currently, laboratory workers are among those most frequently infected (14). Marine mammal strains of Brucella have been reported to cause the infection of a laboratory worker in the UK (15), as well as naturally-acquired infections in Peru (16) and New Zealand (17).Brucella species and biovars, preferential hosts, and pathogenicity for humans are depicted in

Evaluation of an Enzyme-Linked Immunosorbent Assay Using Recombinant Major Surface Protein 5 for Serological Diagnosis of Bovine Anaplasmosis in Venezuela

An indirect enzyme-linked immunosorbent assay (ELISA) was developed for the serological diagnosis of bovine anaplasmosis with purified recombinant major surface protein 5 (MSP5) of Anaplasma marginale produced in Escherichia coli. Serum antibody responses against MSP5 were detected in calves experimentally infected with A. marginale as early as 21 days postinfection and reached maximum titers at 28 days postinfection. The MSP5 ELISA performed with serum samples taken from field cattle from different regions of Venezuela showed a seroprevalence of 47%, which seems to be in accordance with the reported epidemiological status of bovine anaplasmosis in Venezuela. Positive results obtained in the MSP5 ELISA were further confirmed by immunoblotting, with the recombinant MSP5 as the antigen. Thus, these results confirmed the importance of MSP5 as a suitable antigen for the serological diagnosis of bovine anaplasmosis.     

A novel evaluation of World No Tobacco day in Latin America

Background

World No Tobacco Day (WNTD), commemorated annually on May 31, aims to inform the public about tobacco harms. Because tobacco control surveillance is usually annualized, the effectiveness of WNTD remains unexplored into its 25th year.

Objective

To explore the potential of digital surveillance (infoveillance) to evaluate the impacts of WNTD on population awareness of and interest in cessation.

Methods

Health-related news stories and Internet search queries were aggregated to form a continuous and real-time data stream. We monitored daily news coverage of and Internet search queries for cessation in seven Latin American nations from 2006 to 2011.

Results

Cessation news coverage peaked around WNTD, typically increasing 71% (95% confidence interval [CI] 61–81), ranging from 61% in Mexico to 83% in Venezuela. Queries indicative of cessation interest peaked on WNTD, increasing 40% (95% CI 32–48), ranging from 24% in Colombia to 84% in Venezuela. A doubling in cessation news coverage was associated with approximately a 50% increase in cessation queries. To gain a practical perspective, we compared WNTD-related activity with New Year’s Day and several cigarette excise tax increases in Mexico. Cessation queries around WNTD were typically greater than New Year’s Day and approximated a 2.8% (95% CI –0.8 to 6.3) increase in cigarette excise taxes.

Conclusions

This novel evaluation suggests WNTD had a significant impact on popular awareness (media trends) and individual interest (query trends) in smoking cessation. Because WNTD is constantly evolving, our work is also a model for real-time surveillance and potential improvement in WNTD and similar initiatives.     

Safety of the blood supply in Latin America

Appropriate selection of donors, use of sensitive screening tests, and the application of a mandatory quality assurance system are essential to maintain the safety of the blood supply. Laws, decrees, norms, and/or regulations covering most of these aspects of blood transfusion exist in 16 of the 17 countries in Latin America that are the subject of this review. In 17 countries, there is an information system that, although still incomplete (there are no official reports on adverse events and incidents), allows us to establish progress made on the status of the blood supply since 1993. Most advances originated in increased screening coverage for infectious diseases and better quality assurance. However, in 2001 to 2002, tainted blood may have caused infections in 12 of the 17 countries; no country reached the number of donors considered adequate, i.e., 5% of the population, to avoid blood shortages, or decreased significantly the number of blood banks, although larger blood banks are more efficient and take advantage of economies of scale. In those years, paid donors still existed in four countries and replacement donors made up >75% of the blood donors in another eight countries. In addition, countries did not report the number of voluntary donors who were repeat donors, i.e., the healthiest category. In spite of progress made, more improvements are needed.     

Highly Sensitive Enzyme Immunoassays for the Detection of β-Lactam Antibiotics

Specific and sensitive antibodies against β-lactam antibiotics are difficult to raise due to the chemical reactivity of the β-lactam ring. The antibiotic-protein conjugates used as the immunogen can easily react with primary amino groups as for instance the ε-amino group of lysine side chains in proteins. This leads to the degradation of the immunogen to a complex mixture and, therefore, to an unpredictable immune response of the host animal. We produced antisera against the hydrolyzed form of β-lactam antibiotics by immunizing rabbits with stable conjugates mimicking this form. Addition of penicillinase in the immunoassay leads to hydrolysis of the β-lactam antibiotic, which is recognized by the antiserum. In a competitive enzyme-immunoassay, benzylpenicillin could be detected at levels of 0.05 ng ml -1 and cloxacillin at levels of 0.1 ng ml -1 in pasteurized milk. The sensitivity and also the selectivity of these EIAs are remarkably high and offer a wide range of different applications. The procedure should be applicable for other β-lactam antibiotics, and may therefore in future play an important role in food quality control and assurance.