Enzyme-linked Immunosorbent Assay Test Kit Research Articles

Deoxynivalenol and its conjugates in beer: A critical assessment of data obtained by enzyme-linked immunosorbent assay and liquid chromatography coupled to tandem mass spectrometry

Enzyme-linked immunosorbent assays (ELISAs) are often employed for the control of deoxynivalenol (DON) in barley and other intermediates involved in beer production chain. Because of the occurrence of high levels of DON-3-glucoside (DON-3-Glc) in malt and beer that have been reported for the first time in our earlier study, research focused on the accuracy of DON determination by immunoassays in cereal-based matrices has been initiated. DON-3-Glc was strongly cross-reacting in all examined commercial ELISA test kits (Ridascreen ® DON (R-Biopharm), Veratox 5/5 DON ® (Neogen Corporation), Deoxynivalenol EIA (Euro-Diagnostica), and AgraQuant ® DON Assay 0.25/5.0 Test Kit (Romer Labs). The highest overestimation in beer analysis, up to 1000%, when taking the DON content determined by liquid chromatography coupled to tandem mass spectrometry (LC–MS/MS) as a reference method, was obtained by AgraQuant assay. Besides of DON-3-Glc and 3- and 15-acetyldeoxynivalenol (ADONs), also other, not known yet, matrix components contributed to false positive results. Similar phenomenon, although in a lesser extent due to lower content of these substances, was observed for using ELISA in the analysis of wheat. The relationship between a way of sample handling and DON overestimation was demonstrated; higher ELISA response was measured in an aqueous extract compared to that prepared by acetonitrile–water (84:16, v/v). Most of cross-reacting co-extracts were removed by MycoSep™# 226 cartridge, what leads us to the hypothesis on the presence of currently unknown cross-reactive species.

Detection of Melamine Using Commercial Enzyme-Linked Immunosorbent Assay Technology

Recent cases of adulteration with melamine have led to the need for rapid and reliable screening methods. To meet this need, commercial enzyme-linked immunosorbent assay (ELISA) test kits for the detection of triazines were evaluated. The recently released Melamine Plate kit (Abraxis, Warminster, Pa.) displayed a limit of detection of 9 ng/ml for melamine in phosphate-buffered saline (PBS) and approximately 1 μg/ml for melamine added to dog food. An atrazine ELISA test kit produced by Abraxis required 0.2 mg/ml to generate a response more than four times the standard deviation from background. In contrast, with the EnviroGard Triazine Plate kit (Strategic Diagnostics, Inc., Newark, Del.), 1.5 mg/ml melamine in PBS generated a signal only one standard deviation from background, which was insufficient to define a limit of detection. Extraction based on dilution with 105mM sodium phosphate/75mM NaCl/2.5% nonfat milk/0.05% Tween 20 (UD) enabled detection of fivefold less melamine in dog food than did use of the procedure recommended by the manufacturer, which entailed extraction into 60% methanol, sonication, centrifugation, filtration, and further dilution into 10% methanol/PBS. Using the Abraxis Melamine ELISA, both extraction protocols yielded identical results with a dog food sample adulterated with mela-mine. The recovery of melamine spiked into gravy from dog food using UD was 74% ± 4%. In conclusion, the recently released Abraxis ELISA for melamine proved to be a useful alternative to more cumbersome methods.

Application of an enzyme-linked immunosorbent assay for the detection of clenbuterol residues in swine urine and feeds

The present study outlines applications of an enzyme-linked immunosorbent assay (ELISA) for the analysis of clenbuterol residues. Antisera were raised from rabbits immunized with diazotized clenbuterol-bovine serum albumin (BSA) conjugate. The assay was specific to clenbuterol with a half-maximum inhibition concentration (IC50) of 1.8 ng/mL and 2.5 ng/mL in blank swine urine and phosphate buffer solution, respectively. The assay had high cross-reactivity (86%) with mabuterol, but low with other adrenergic agonists and antagonists. The average recovery of clenbuterol, as measured with the ELISA, ranged from 90% to 112% in swine urine samples and from 86% to 95% in feeds, respectively. This new assay was compared with commercial ELISA test kits. An excellent correlation (r 2 = 0.98) between the two methods and satisfactory recoveries suggest that the new assay can be suitable for the determination of clenbuterol residues in real samples. The assay was used to analyze clenbuterol residues in 103 swine urine samples and 68 feed samples collected from northern China. Approximately 50% of the urine samples and 25% of the feed samples analyzed were found positive (concentration of clenbuterol ≥ 1 ppb). The results indicate that clenbuterol was misused in some of the areas surveyed.

Assessment of Two Enzyme-Linked Immunosorbent Assay Tests Marketed for Detection of Ruminant Proteins in Finished Feed

The performance characteristics of two enzyme-linked immunosorbent assay (ELISA) test kits, ELISA Technologies' MELISA-Tek test and Tepnel BioSystems' BioKit for (Cooked) Species Identification test, designed to detect ruminant proteins in animal feed, were evaluated. The test kits were evaluated by using acceptance criteria developed by the U.S. Food and Drug Administration's Center for Veterinary Medicine Office of Research for evaluating selectivity, sensitivity, ruggedness, and specificity. The acceptance criteria for determining success used a statistical approach requiring a 90% probability of achieving the correct response within a 95% confidence interval. In practice, this measure requires the test to achieve the correct response 58 times for every 60 samples evaluated, or a 96.7% accuracy rate. A minimum detection level of 0.1% bovine meat and bone meal (BMBM) was required, consistent with the sensitivity of the analytical methods presently used by the U.S. Food and Drug Administration. Selectivity was assessed by testing 60 dairy feed samples that contained no added animal proteins; sensitivity was determined by evaluating 60 samples (per level of fortification) of this same feed that contained 0.025, 0.05, 0.1, 0.25, 0.5, 1, or 2% BMBM. The MELISA-Tek test passed the acceptance set-point criteria for selectivity assessment but failed the sensitivity assessment at all levels except at the 2% level. The MELISA-Tek test came close to passing at the 1% level, detecting true-positive findings at a rate of 93%, but failed at lower levels, in spite of the label claim of 0.5% sensitivity. The BioKit for (Cooked) Species Identification test detected only 2 of 17 samples fortified at the 2% BMBM level and failed to detect any other BMBM-fortified samples. The results of this evaluation indicate that neither test is adequate for regulatory use.

A new ELISA kit which uses a combination of Plasmodium falciparum extract and recombinant Plasmodium vivax antigens as an alternative to IFAT for detection of malaria antibodies

BackgroundThe methods most commonly used to measure malarial antibody titres are the Indirect Fluorescence Antibody Test (IFAT), regarded as the gold standard, and the Enzyme-Linked ImmunoSorbent Assay (ELISA). The objective here was to assess the diagnostic performance, i.e. the sensitivity and specificity, of a new malaria antibody ELISA kit in comparison to IFAT. This new ELISA kit, the ELISA malaria antibody test (DiaMed), uses a combination of crude soluble Plasmodium falciparum extract and recombinant Plasmodium vivax antigens.MethodsTwo groups were used: 95 samples from malaria patients to assess the clinical sensitivity and 2,152 samples from blood donors, who had not been exposed to malaria, to assess the clinical specificity.ResultsThe DiaMed ELISA test kit had a clinical sensitivity of 84.2% and a clinical specificity of 99.6% as compared with 70.5% and 99.6% respectively, using the IFAT method. The ELISA method was more sensitive than the IFAT method for P. vivax infections (75% vs. 25%). However, in 923 malaria risk donors the analytical sensitivity of the ELISA test was 40% and its specificity 98.3%, performances impaired by large numbers of equivocal results non-concordant between ELISA and IFAT. When the overall analytical performances of ELISA was compared to IFAT, the ELISA efficiency J index was 0.84 versus 0.71 for IFAT. Overall analytical sensitivity was 93.1% and the analytical specificity 96.7%. Overall agreement between the two methods reached 0.97 with a reliability k index of 0.64.ConclusionThe DiaMed ELISA test kit shows a good correlation with IFAT for analytical and clinical parameters. It may be an interesting method to replace the IFAT especially in blood banks, but further extensive investigations are needed to examine the analytical performance of the assay, especially in a blood bank setting.

Normal levels of ADAMTS13 and factor H are present in the pharmaceutically licensed plasma for transfusion (Octaplas®) and in the universally applicable plasma (Uniplas) in development

The pathomechanism of thrombotic thrombocytopenic purpura (TTP) and atypical haemolytic uraemic syndrome (aHUS) is associated with a severe deficiency of ADAMTS13 and factor H. The aim of this study was to quantify the levels of ADAMTS13 and factor H in the pharmaceutically licensed plasma for transfusion, Octaplas, and the universally applicable plasma, Uniplas (development product, working title). Furthermore, Octaplas batches of blood groups A, B, O, AB, and plasmas derived from different sources were compared. Twenty-four Octaplas and three Uniplas batches were selected for the study. ADAMTS13 activities were measured by fluorescence resonance energy transfer assay, ADAMTS13 antigen levels were quantified using enzyme-linked immunosorbent assay test kit, while factor H antigen levels were detected using radial immunodiffusion (RID) methods. In addition, von Willebrand factor (vWF) multimeric analyses were performed. Both Octaplas, produced from US and European plasma of different blood groups, and Uniplas contain ADAMTS13 antigen and activity levels as well as factor H concentrations at normal levels without significant differences. In addition, Octaplas and Uniplas show a vWF multimeric pattern comparable to normal plasma. The study revealed that Octaplas and Uniplas contain normal levels of ADAMTS13 at low batch-to-batch variations. Therefore, both products can substitute the missing or neutralized protease activity in TTP patients and thereby limit vWF-dependent (platelet-related) thrombosis. In addition, both plasma products contain factor H at a physiological level, and, thus can be used efficiently in the treatment of aHUS patients, which have been shown to benefit from plasma administration.

Investigation on sequential extraction of peanut allergens for subsequent analysis by ELISA and 2D gel electrophoresis

A two-step sequential extraction method of peanut proteins was proposed with the aim to investigate the protein composition and allergen content of peanut samples. The extraction procedure reported is fully compatible with subsequent analysis by enzyme-linked immunosorbent assays (ELISA) as well as 2D gel electrophoresis (2D PAGE). This sequential extraction method was used to study three different peanut varieties and three different types of food processing. Peanuts were analysed for total protein content and the extraction efficiency of raw and processed peanuts was determined. The total protein content of the three peanut varieties was found to be comparable, but their extraction efficiency varies. The peanut extracts were characterised by employing three different ELISA test kits specific to either the allergens Ara h 1 or Ara h 2, or to soluble peanut proteins. The content of both Ara h 1 and Ara h 2 differed in the raw peanut extracts of the three varieties. However, thermal processing resulted in much larger changes in detectability. Blanching significantly increases the detectability of Ara h 2, whereas Ara h 1 detection remains almost unchanged. After roasting a clear decrease of detectability was observed for both Ara h 1 and Ara h 2, although the effect is more severe for Ara h 1. 2D PAGE was employed to compare the protein profiles and abundances of peanut extracts. Statistically relevant differences were observed for the two different protein fractions obtained by using the described method, showing the relevance of this two-step sequential extraction method.

Use of Age and Milk Production Data to Improve the Ability of Enzyme-Linked Immunosorbent Assay Test Results to PredictMycobacterium Aviumssp.ParatuberculosisFecal Culture Status

Cows from 2 California dairies were tested for paratuberculosis at the end of lactation by using fecal culture and a commercially available serum enzyme-linked immunosorbent assay (ELISA) test kit. Individual cow characteristics and production variables were evaluated along with ELISA testing results as predictors of fecal culture status. In multivariable logistic regression analysis, age and a herd-standardized version of 305-day mature equivalent (305 ME) milk production were significant predictors of fecal culture status after adjusting for herd, quarter of the study year, and ELISA sample-to-positive (S/P) ratio. The area under a nonparametric receiver operating characteristic curve was significantly greater for a multivariable model that included age and the level of milk production when compared with a model without these covariates. In conclusion, consideration of cow-level covariates was useful as an aid in predicting Mycobacterium avium ssp. paratuberculosis (MAP) fecal culture status. For a given ELISA S/P ratio, older cows and those with lower 305 ME milk production relative to other cows in the herd were significantly more likely to be shedding MAP in their feces at the end of lactation.

2004 National Atrazine Occurrence Monitoring Program Using the Abraxis ELISA Method

The goal of this project was to gain a better understanding of atrazine occurrence in the United States by surveying drinking water utilities' sources and finished water for atrazine on a weekly basis for seven months. Atrazine is a contaminant of interest because the United States Environmental Protection Agency (USEPA) has found short-term atrazine exposure above the drinking water maximum contaminant level (MCL) to potentially cause heart, lung, and kidney congestion, low blood pressure, muscle spasms, weight loss, and damage to the adrenal glands. Long-term exposure to atrazine concentrations above the drinking water MCL has been linked to weight loss, cardiovascular damage, retinal and muscle degeneration, and cancer. This survey effort improved upon previously conducted atrazine surveys through intensive, high frequency sampling (participating plants sampled their raw and finished water on a weekly basis for approximately seven months). Such an intensive effort allowed the authors to gain a better understanding of short-term atrazine occurrence and its variability in drinking water sources. This information can benefit the drinking water industry by facilitating (1) better atrazine occurrence management (i.e., awareness when plants may be more susceptible to atrazine), (2) more efficient atrazine control (e.g., effective treatment alternatives and more effective response to atrazine occurrence), and (3) treatment cost reduction (e.g., efficient atrazine control can result in substantial cost savings). Forty-seven drinking watertreatment plants located primarily in the Midwestern United States participated in the survey and sampled their raw and finished water on a weekly basis from March through October. Samples were analyzed using the Abraxis enzyme-linked immunosorbent assay (ELISA) test kit. Confirmation samples for quality assurance/quality control (QA/QC) purposes were analyzed using solid-phase extraction (SPE) followed by gas chromatography mass spectrophotometry (GC/MS). Several important conclusions can be drawn from this study including (1) surface waters were confirmed to be more vulnerable to atrazine contamination than groundwater sources, (2) peak atrazine concentrations corresponded well to precipitation/runoff events, and (3) atrazine occurrence tended to be uniform geographically when compared by river drainage basins. In addition, this project confirmed that the Abraxis atrazine ELISA test kit tended to have a positive bias (i.e., the measured ELISA concentration was higher than the actual concentration) in most measured samples. Finished samples tended to have more of a positive bias than raw water samples. Therefore, this bias may limit the effectiveness for ELISA for regulatory monitoring. There are many other applications for ELISA, however, including frequent monitoring for early detections of atrazine concentration changes that might trigger conventional analysis by GC/MS or be used for activated carbon dosing or other treatment operating controls.

Accuracy and Interferences for Enzyme‐Linked Immunoassay Tests for Atrazine

Enzyme‐linked immunosorbent assay (ELISA) methods have become a common alternative to gas chromatography methods for analyzing atrazine in raw and finished drinking water. This study investigated several leading ELISA atrazine test kits currently in use. This work demonstrates that although all the kits analyzed provided precise results, their accuracy varies greatly between kits and water matrixes. In most cases, the ELISA test kits provided positively biased results, although some negatively biased results were also observed. That is, the ELISA atrazine concentrations were usually greater than the actual concentrations. Although a part of this positive bias was because of cross‐reactivity with other chloro‐s‐triazines in the water matrix, much of the observed interference appeared to be related to natural organic matter, chlorination and/or quenching, and other factors. Even though the observed positive bias tends to cause the kits to provide a conservative result with respect to atrazine concentration, the role of ELISA test kits for regulatory purposes requires more study and discussion.

Effect of roasting history and buffer composition on peanut protein extraction efficiency

Peanut is a major allergenic food. Undeclared peanut (allergens) from mis-formulation or contamination during food processing pose a potential risk for sensitized individuals and must be avoided. Reliable detection and quantification methods for food allergens are necessary in order to ensure compliance with food labelling and to improve consumer protection. The extraction of proteins from allergenic foods and complex food products is an important step in any allergen detection method. In this study, the protein extraction efficiency of various buffers prepared in-house and some extraction buffers included in some commercial allergen enzyme-linked immunosorbent assay (ELISA) test kits for peanut determination in food products were tested. In addition, the effect of roasting history on the extractability of peanut protein was investigated by the biuret and the bicinchoninic acid (BCA) assays. Elevated roasting temperatures in food processing were found to have a major impact on protein extraction efficiency by reducing protein yields of oil and dry roasted peanuts by 50-75% and 75-80%, respectively, compared with the raw material. Extraction buffers operating in the higher pH range (pH 8-11) showed best yields.

Immunogenicity of venoms from four common snakes in the South of Vietnam and development of ELISA kit for venom detection

The antigenicity and antigenic relationship between venoms of four common snakes in the South of Vietnam— Trimeresurus popeorum, Calloselasma rhodostoma, Naja naja and Ophiophagus hannah—were studied. Most of venom components expressed antigenicity and produced high titre antivenoms. The venoms share common components and antivenoms cross-reacted along them. Furthermore, cross-reactions were observed among non-common antigens, indicating that they share common epitopes. Hence, using single component as immunogen for species diagnosis of snakebites can reduce cross-reaction, perhaps may not be totally specific. A three-step affinity purification protocol was set up for preparation of species-specific antivenom antibodies. The steps involved affinity chromatography of IgG from hyper-immunized rabbit sera with protein A columns, immuno-affinity chromatography of monovalent antivenom antibodies with respective homologous venom columns, and immuno-absorption of cross-species reacting antibody molecules with heterologous venom columns. The antibodies were then used for construction of an enzyme-linked immunosorbent assay (ELISA) test kit. The kit can differentiate among the four common snake venoms in various types of samples with the detection limit of 0.2–1.6 ng/ml, depending on the type of samples and species of the snake. The efficacy of this kit for snake venom detection was successfully demonstrated in experimental envenomation in rats. Preliminary evaluation with 140 samples taken from 88 human snakebite victims in Vietnam showed that the kit could detect venom in human samples and would be a very useful tool for fast identification of snakebites in clinics.

The seroprevalence of Johne's disease in Georgia beef and dairy cull cattle.

Beef and dairy cattle serum samples, collected during 2000 at sale barns throughout Georgia, were obtained from the Georgia State Brucellosis Laboratory and were used to conduct a retrospective epidemiological study. Statistical samplings of 5,307 sera, from over 200,000 sera, were tested for antibodies to Mycobacterium avium ssp. paratuberculosis, (Johne's disease) using a commercial enzyme-linked immunosorbent assay test kit. An overall period seroprevalence in all classes of cattle tested was 4.73%. The period seroprevalence in dairy cattle was 9.58%, in beef cattle it was 3.95%, and in cattle of unknown breed it was 4.72%. It was concluded that the seroprevalence of Johne's disease in cull beef and dairy cattle in Georgia is economically significant.

Determination of Rendering Plant Sterilization Conditions Using a Commercially Available ELISA Test Kit Developed for Detection of Cooked Beef

A commercially available enzyme-linked immunosorbent assay (ELISA) test kit developed for detection of cooked beef in meat samples was used to determine appropriate heat treatment of rendered materials. An improved extraction procedure increased the absolute difference in R-values between 2 rendered materials treated under different conditions (average temperature 129 and 134 degrees C, respectively). To evaluate the influence of the main sterilization parameters on ELISA results, a factorial design approach was used. The parameters investigated were temperature, time, particle size, and meat composition. Lean meat samples containing beef and pork were sterilized under strictly controlled conditions in a laboratory autoclave. The experiments demonstrated that the R-values obtained with the ELISA test kit for beef are strongly influenced by temperature and time, whereas particle size has a minor influence. The proportion of bovine material did not have any impact on R-values. Autoclave-processed lean meat samples were analyzed by using an ELISA test kit for pork, which was validated in a collaborative trial. The ELISA test kit for pork proved to be more sensitive for the investigated parameters, thus verifying and extending previous investigations.

False-positive urine β-HCG in a woman with a tubo-ovarian abscess

Modern urine β-human chorionic gonadotropin (HCG) assays that use enzyme-linked immunosorbent assay (ELISA) technology are sensitive and specific for diagnosing pregnancy, both intrauterine and ectopic, and have become indispensable to the practice of Emergency Medicine. A urine HCG test is often relied on by the Emergency Physician as a critical component in the diagnostic regimen of a patient with a possible ectopic pregnancy. We report a case of a false-positive urine β-HCG test in a patient with a ruptured tubo-ovarian abscess. Though false-positive pregnancy tests with tubo-ovarian abscesses have previously been reported with older methods of HCG detection, we believe that this is the first case where the pregnancy test was the modern ELISA type. The mechanism for the false-positive reaction in this case is unknown, but time may show that the ELISA test kit, like its predecessors, may occasionally give a false-positive reaction in this class of patients.

Method of Determination of Appropriate Heat Treatment of Animal Meal by Immunoassay Developed for Detection of Cooked Beef: Interlaboratory Study

An interlaboratory trial was conducted for the validation of an enzyme-linked immunosorbent assay (ELISA) method for determination of appropriate heat treatment of animal meal. A commercially available ELISA test kit developed for the identification of beef in cooked food was used in the study. Twelve laboratories from 7 European countries examined 2 different analytical protocols to establish the most appropriate analytical method. Three different samples were used, 2 animal waste materials sterilized at 129 and 134 degrees C (wet conditions), respectively, and a meat and bone meal material processed at dry conditions (maximum temperature, 140 degrees C). Statistical evaluation applying t-statistics showed that the animal meal treated according to European legislation (>133 degrees C) was clearly distinguishable from the 2 other test materials at a 99% confidence level using both analytical protocols. This method can be considered as a complementary test to the immunoassay developed for the detection of pork in cooked food that is already applied in routine analysis for the surveillance of rendering plants.

European brown hare syndrome in free-ranging European brown and mountain hares from Switzerland.

From 1997 to 2000, complete necropsy and histopathologic investigations were performed on 157 free-ranging European brown hares (Lepus europaeus) found dead throughout Switzerland. Organ samples of all these individuals (157 livers and 107 spleens available) were tested for European brown hare syndrome virus (EBHSV)-antigen by enzyme-linked immunosorbent assay (ELISA) test kit. Furthermore, 60 additional blood samples were tested for antibodies against EBHSV by ELISA. In addition, liver samples of 87 free-ranging mountain hares (Lepus timidus) hunted in 1996 were tested for EBHSV-antigen. In two European brown hares from southern Switzerland lesions suggestive of changes induced by EBHSV were present, and high titers of EBHSV-antigen were detected in both liver and spleen samples of these animals. Based on negative staining electron microscopy investigations of liver and spleen homogenates, we observed calicivirus in one antigen-positive hare. Low EBHSV-antigen titers were found in three additional European brown hares from central and western Switzerland, but EBHS-lesions were absent. Antibodies against EBHSV were not detected in any of the sera of European brown hares, and EBHSV-antigen was not found in the samples of mountain hares. This is the first report of EBHS in European brown hares from Switzerland.

1998/99 national survey of pesticides in grdundwater using GCMS and ELISA

A total of 95 wells throughout New Zealand were sampled during summer 1998/99 and analysed for a range of pesticides using gas chroma‐tography‐mass spectroscopy (GCMS). Thirty‐three wells (35%) had pesticides including triazine metabolites detected, with 18 wells (19%) having two or more pesticides detected. Only one well (K38/ 0172) had pesticides detected at levels greater than the maximum acceptable value (MAV) for drinking water. There were 20 different pesticides detected, usually at very low concentrations, as well as two triazine metabolites. Fifty‐seven out of the 75 pesticide detections (76%) belonged to the triazine group, with only five of these (9%) being >1 mg m−3 and 74% of these being <0.1 mg m‐3. Three of the 18 non‐triazine pesticide detections (17%) were > 1 mg m−3 with all three detections being in Well K38/ 0172. About half (44%) of the non‐triazine pesticides were <0.1 mg M−3. Apart from Well K38/0172, the highest levels of pesticides with respect to the MAV were seen for simazine and terbuthylazine, which were 16 and 44% of the MAV, respectively, with the remaining pesticides all below 4% of their MAV. This indicates that, although there were very low levels of pesticides present in 35% of the wells, there would be no significant health risk based on the pesticides analysed from drinking the groundwater investigated with the exception of Well K38/0172. There was a significant decrease in the detection limits for many pesticides for the 1998/99 survey compared to the two earlier surveys. When the detection limits for the earlier surveys were applied to the present survey there would have been a total of 10 wells out of the 95 sampled (11%) with pesticides detected. This compared with 7% of the 82 wells in 1990 and 13.6% of the 116 wells in 1994 with detectable pesticides, indicating that a similar percentage of wells have had detectable pesticides in each survey once correction for variable detection limits has been made. There was a significant difference between wells with and without pesticide detections for the following factors: wells with pesticide detections had shallower well screens, were screened nearer the water table, had lower pH values and higher nitrate levels. Samples for analysis using two enzyme‐linked immunosorbent assay (ELISA) atrazine test kits were collected at the same time as the other pesticide samples. The “normal” atrazine ELISA test kit only showed four positive detections, mainly because of a relatively high detection limit. The high sensitivity (HS) atrazine test kits detected pesticides in 20 of the 28 samples that were tested. Nineteen of the 20 detections had pesticide detections using GCMS, with the other eight samples having no pesticides detected using GCMS. The HS atrazine ELISA test kit appears to be a useful indicator of pesticide contamination for New Zealand groundwater systems at a relatively low level of detection. All the positive GCMS results that were tested gave positive test kit results and there was only one potential false positive result. This is probably because the majority of pesticides detected in groundwater in New Zealand are triazines, and the HS atrazine kit has cross reactivity with most triazines and related pesticides.

ANALYSIS OF PESTICIDE RUNOFF FROM MID-TEXAS ESTUARIES AND RISK ASSESSMENT IMPLICATIONS FOR MARINE PHYTOPLANKTON

During 1993, estuarine surface water samples were collected from the mid-Texas coast (Corpus Christi to Port Lavaca, TX). Agricultural watershed areas as well as tidal creeks immediately downstream were chosen as sampling sites along with adjoining bay sampling stations. Collections were made throughout the growing season (February to October 1993) before and after periods of significant (>1.25 cm) rainfall. All samples were initially screened for the presence of pesticides using enzyme-linked immunosorbent assay (ELISA) test kits (EnviroGard™) for triazine herbicides and carbamate insecticides. All samples were extracted and then analyzed using gas chromatography (GC) for quantification of atrazine. Only samples testing positive for carbamate insecticides via ELISA were further extracted for GC analysis to quantify aldicarb and carbofuran. Additionally, laboratory toxicity tests using phytoplankton were examined from published, peer-reviewed literature and compared with the atrazine field levels found in Texas. Results of ELISA screening indicated the presence of triazine herbicides in nearly all samples (>93%). GC analysis further confirmed the presence of atrazine concentrations ranging from <0.01–62.5 μg/L. Screening tests also found detectable levels of carbamate insecticides (aldicarb and carbofuran) that were also confirmed and quantified by GC. Comparison of measured concentrations of atrazine compared with published toxicity tests results indicated that there was a potential environmental risk for marine/estuarine phytoplankton in surface waters of Texas estuaries, particularly when the chronic nature of atrazine exposure is considered.

Comparison of a Clinic-Based ELISA Test Kit with the Immunofluorescence Test for the Assay of Ehrlichia Canis Antibodies in Dogs

The "gold standard" for the detection of antibodies to Ehrlichia canis, the cause of canine monocytic ehrlichiosis (CME), is the indirect immunofluorescence antibody (IFA) test. The IFA test however is generally available only in selected laboratories and requires extensive equipment and trained personnel. A double-blind study was conducted to compare the ability of an in-clinic standardized enzyme-linked immunosorbent assay (ELISA) test kit to measure E. canis IgG antibodies in dogs compared with the standard IFA technique. A good correlation was found between the 2 techniques (r2 = 0.8793; P < 0.0001). Evidence for the sensitivity of the ELISA technique for the early detection of E. canis IgG antibodies was demonstrated by comparing the appearance of E. canis antibody titers by the IFA and ELISA techniques after artificial infection of 2 sets of dogs. In both experimental infections, both tests were equally sensitive for the early detection of IgG antibodies against E. canis, and the results correlated well with the appearance of fever and clinical signs. Proposed application of the in-clinic ELISA test is to aid in the diagnosis of CME.