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1.
Pancreatitis is recognized as an important cause for morbidity and mortality in cats, but diagnosis remains difficult in many cases. As a first step in trying to identify a better diagnostic tool for feline pancreatitis the objective of this project was to develop and analytically validate a radioimmunoassay for the measurement of feline pancreatic lipase immunoreactivity (fPLI). Feline pancreatic lipase (fPL) was purified from pancreatic tissue and antiserum against fPL was raised in rabbits. Tracer was produced by iodination of fPL using the chloramine T method. A radioimmunoassay was established and analytically validated by determination of sensitivity, dilutional parallelism, spiking recovery, intra-assay variability, and interassay variability. A control range for fPLI in cat serum was established from 30 healthy cats using the central 95th percentile. The sensitivity of the assay was 1.2 microg/L. Observed to expected ratios for serial dilutions ranged from 98.8% to 164.3% for 3 different serum samples. Observed to expected ratios for spiking recovery ranged from 76.9% to 147.6% for 3 different serum samples. Coefficients of variation for intra- and interassay variability for 4 different serum samples were 10.1%, 4.5%, 2.2%, and 3.9% and 24.4%, 15.8%, 16.6%, and 21.3%, respectively. A reference range for fPLI was established as 1.2 to 3.8 microg/L. We conclude that the assay described is sensitive, accurate, and precise with limited linearity in the lower and limited reproducibility in the lower and higher end of the working range. Further studies to evaluate the clinical usefulness of this assay are needed and in progress.  相似文献   

2.
OBJECTIVE: To develop and validate a radioimmunoassay (RIA) for measuring canine pancreatic lipase immunoreactivity (cPLI) in serum obtained from dogs. SAMPLE POPULATION: Serum samples from 47 healthy dogs. PROCEDURES: Canine pancreatic lipase (cPL) was purified from pancreatic specimens of dogs. Antibodies against cPL were raised in rabbits and purified by use of affinity chromatography. A tracer was produced by iodination of cPL with 125I. An RIA was established and validated by determination of sensitivity, working range, dilutional parallelism, spiking recovery, and intra- and interassay variability. A reference range for cPLI in serum was established by use of the central 95th percentile for samples obtained from 47 healthy dogs. RESULTS: Sensitivity and upper limit of the working range were 0.88 and 863 microg/L, respectively. Observed-to-expected ratios for serial dilutions ranged from 84.9 to 116.5% for 4 samples. Observed-to-expected ratios for spiking recovery ranged from 82.8 to 128.6% for 4 samples. Coefficients of variation for intra-assay variability for 4 serum samples were 18.3, 4.2, 3.5, and 8.9%, whereas interassay coefficients of variation were 29.2, 6.2, 3.9, and 4.4%, respectively. The reference range was 4.4 to 276.1 microg/L. CONCLUSIONS AND CLINICAL RELEVANCE: We conclude that the RIA described is sensitive, linear, accurate, precise, and reproducible, with limited accuracy in the high end of the working range and limited precision and reproducibility in the low end of the working range. Additional studies are needed to evaluate whether this degree of accuracy, precision, and reproducibility will negatively impact clinical use of this assay.  相似文献   

3.
OBJECTIVE: To determine cellular immunolocalization of canine gastric lipase (cGL) and canine pancreatic lipase (cPL) in various tissues obtained from clinically healthy dogs. SAMPLE POPULATION: Samples of 38 tissues collected from 2 climically healthy dogs. PROCEDURES: The cGL and cPL were purified from gastric and pancreatic tissue, respectively, obtained from dogs. Antisera against both proteins were developed, using rabbits, and polyclonal antibodies were purified by use of affinity chromatography. Various tissues were collected from 2 healthy dogs. Primary antibodies were used to evaluate histologic specificity. Replicate sections from the collected tissues were immunolabeled for cGL and cPL and examined by use of light microscopy. RESULTS: Mucous neck cells and mucous pit cells of gastric glands had positive labeling for cGL, whereas other tissues did not immunoreact with cGL. Pancreatic acinar cells had positive labeling for cPL, whereas other tissues did not immunoreact with cPL. CONCLUSIONS AND CLINICAL RELEVANCE: We concluded that cGL and cPL are exclusively expressed in gastric glands and pancreatic acinar cells, respectively. Also, evidence for cross-immunoreactivity with other lipases or related proteins expressed by other tissues was not found for either protein. Analysis of these data suggests that gastric lipase is a specific marker for gastric glands and that pancreatic lipase is a specific marker for pancreatic acinar cells. These markers may have clinical use in the diagnosis of gastric and exocrine pancreatic disorders, respectively.  相似文献   

4.
Recently, a radioimmunoassay (RIA) for measurement of canine pancreatic lipase immunoreactivity (cPLI) in serum was developed and validated. However, RIAs require frequent use of radioactive materials. Therefore, the goal of this project was to develop and validate an enzyme-linked immunosorbent assay (ELISA) for cPLI. After purifying cPL, we developed and purified antiserum against cPL in rabbits. The purified antibody was bound to microtitre plates and used to capture antigen. A portion of the purified antibody was biotinylated and used to identify the captured antigen. Streptavidin labelled with horseradish peroxidase and a horseradish peroxidase substrate were used for detection. The assay was validated by determination of sensitivity, working range, linearity, accuracy, precision, and reproducibility. The reference interval for serum cPLI was determined by the central 95th percentile in 74 clinically healthy dogs: 2.2 to 102.1 μg/L. The sensitivity and the upper limit of the working range were 0.1 and 999.2 μg/L, respectively. The ratios of observed to expected values for dilutional parallelism for 6 serum samples ranged from 0.0 to 148.8%; the ratios for spiking recovery for 4 serum samples ranged from 90.4 to 112.6%, assuming 55% recovery of the cPL. Coefficients of variation for intra- and interassay variability for 6 different serum samples were 2.4, 3.4, 4.1, 5.8, 7.4, and 10.0% and 5.9, 7.7, 11.6, 13.9, 23.5, and 46.2%, respectively. We conclude that the ELISA described here is sufficiently sensitive, linear, accurate, precise, and reproducible for clinical application. Evaluation of its clinical usefulness for the diagnosis of exocrine pancreatic disorders in dogs is under way.  相似文献   

5.
OBJECTIVE: To develop and validate an ELISA for measurement of serum canine pepsinogen A (cPG A) as a diagnostic marker of gastric disorders in dogs and to measure serum cPG A in healthy dogs after food deprivation and after feeding. SAMPLE POPULATION: Sera from 72 healthy dogs. PROCEDURE: A sandwich ELISA was developed and validated. The reference range for serum concentrations of cPG A was determined in 64 healthy dogs. Postprandial changes in serum concentrations of cPG A were evaluated in 8 healthy dogs. RESULTS: Assay sensitivity was 18 microg/L, and the maximum detectable concentration was 1,080 microg/L. The observed-to-expected ratio (O:E) for 3 serial dilutions of 3 serum samples ranged from 69.3 to 104.1%. The O:E for 3 serum samples spiked with 8 concentrations of cPG A ranged from 58.8 to 120.4%. Coefficients of variation for intra- and interassay variability of 3 serum samples ranged from 7.6 to 11.9% and from 10.1 to 13.1%, respectively. Mean +/- SD serum concentration of cPG A in healthy dogs was 63.8 +/- 31.0 microg/L and the reference range was 18 to 129 microg/L. Significant increases in serum concentrations of cPG A were observed between 1 and 7 hours after feeding. CONCLUSIONS AND CLINICAL RELEVANCE: The ELISA for measuring cPG A was sufficiently sensitive, linear, accurate, precise, and reproducible for clinical use. Serum concentrations of cPG A increase substantially after feeding, which should be taken into account when conducting clinical studies.  相似文献   

6.
An enzyme-linked immunosorbent assay (ELISA) for canine blood apolipoprotein (apo) B-100 and A-I was developed. The working range for the assay was 1.8 to 28.7 ng/well for apoB-100 and it was 50 to 410 ng/well for apoA-I. The intra- and inter-assay coefficients of variation for the assay for apoB-100 were 5.4 and 6.9%, respectively, and for apoA-I they were 5.8 and 10.6%, respectively. The average concentrations of apoB-100 and A-I in 25 beagles (males, aged 3-4 years) were 0.084 +/- 0.028 (mean +/- SD) mg/ ml and 6.29 +/- 1.55 mg/ml, respectively. The ratios of canine (C) apoB-100 to CapoA-I were 1.41 +/- 0.58%. The respective concentrations in one case of hyperlipidemia with systemic atherosclerosis were 0.454 mg/ml and 11.28 mg/ml (a ratio of 4.03%). These values were larger than those of the controls. These results suggest that the measurements of CapoB-100 and A-I concentrations by this newly developed ELISA are helpful for diagnosis of lipidosis.  相似文献   

7.
An automated colorimetric method for determining lipase activity in canine sera was evaluated for precision, linearity and correlation to existing assay methods. The colorimetric method was a commercial reagent that used a series of enzymatic reactions based on the hydrolysis of 1,2 diglyceride by pancreatic lipase. Within-run and between-run coefficients of variation were < 6.8% and < 8.3%, respectively. Linearity was determined to be at least 1366 U/L. Canine serum lipase concentrations attained using the colorimetric method were compared to both titrimetric and dry-film methods for measuring serum lipase activity, resulting in significant (P < or = 0.05) correlation coefficients of 0.92 and 0.77, respectively. Canine serum lipase concentrations measured using the colorimetric assay on 2 different automated analyzers had a significant (P < or = 0.05) correlation coefficient of 0.92. A laboratory reference range using serum samples from 56 healthy dogs (0-561 U/L) was established. There were no significant (P < or = 0.05) differences in mean serum lipase concentrations comparing male and female dogs or comparing young dogs (< or = 3 y) to mature (4-7 y) and older (> 7 y) dogs using this assay. It was concluded that the automated colorimetric assay was a reliable indicator of canine serum lipase activity and offered several advantages, including small sample volume and short analysis time.  相似文献   

8.
OBJECTIVE: To purify neutrophil elastase (NE) from dog blood and develop and validate an ELISA for the measurement of canine NE (cNE) in canine serum as a marker for gastrointestinal tract inflammation. SAMPLE POPULATION: Neutrophils from 6 dogs immediately after they were euthanatized and serum from 54 healthy dogs. PROCEDURES: cNE was purified from blood by use of dextran sedimentation, repeated cycles of freezing-thawing and sonication, cation-exchange chromatography, and continuous elution electrophoresis. Antibodies against cNE were generated in rabbits, and an ELISA was developed and validated by determination of sensitivity, dilutional parallelism, spiking recovery, intra-assay variability, and interassay variability. A reference range was established by assaying serum samples from the 54 healthy dogs and by use of the lower 97.5th percentile. RESULTS: cNE was successfully purified from blood, and antibodies were successfully generated in rabbits. An ELISA was developed with a sensitivity of 1,100 mug/L. The reference range was established as < 2,239 mug/L. Ratios of observed-to-expected results for dilutional parallelism for 4 serum samples ranged from 85.4% to 123.1%. Accuracy, as determined by spiking recovery, ranged from 27.1% to 114.0%. Coefficient of variation for 4 serum samples was 14.2%, 16.0%, 16.8%, and 13.4%, respectively, for intra-assay variability and 15.4%, 15.0%, 10.5%, and 14.6%, respectively, for interassay variability. CONCLUSIONS AND CLINICAL RELEVANCE: The purification protocol used here resulted in rapid and reproducible purification of cNE with a high yield. The novel ELISA yielded linear results and was accurate and precise. Additional studies are needed to evaluate the clinical usefulness of this assay.  相似文献   

9.
OBJECTIVE: To determine the characteristics of an automated canine C-reactive protein (CRP) assay and evaluate 2 human CRP assays for use in dogs. Animals-56 client-owned dogs with pyometra and 11 healthy control dogs. PROCEDURES: Samples from 11 dogs with high (> 100 mg/L) or low (< 10 mg/L) CRP concentrations (determined by use of a canine ELISA) were evaluated by use of the automated canine CRP assay. Intra- and interassay imprecision was determined (by use of those 2 plasma pools), and assay inaccuracy was assessed by use of logistic regression analysis of results obtained via ELISA and the automated canine CRP assay. Two automated human CRP assays were used to measure plasma CRP concentration in 10 dogs. RESULTS: By use of the ELISA, mean +/- SD plasma CRP concentration was 96.1 +/- 38.5 mg/L and 10.1 +/- 23.2 mg/L in dogs with pyometra and control dogs, respectively. The automated canine assay had intra-assay coefficients of variation (CVs) of 7.8% and 7.9%, respectively, and interassay CVs of 11.1% and 13.1%, respectively. Results from the automated assay were highly correlated with results obtained via ELISA. The human assay results did not exceed 0.4 mg/L in any dog. CONCLUSIONS AND CLINICAL RELEVANCE: The automated canine CRP assay had less interassay imprecision, compared with the ELISA. The 2 human CRP assays were not suitable for analysis of canine plasma samples. The automated canine CRP assay was more precise than the ELISA for serial evaluations of plasma CRP concentration in dogs.  相似文献   

10.
OBJECTIVE: To assess the safety of endoscopic retrograde pancreatography (ERP) in dogs by performing repeated clinical examinations and laboratory analyses of serum amylase, lipase, canine trypsin-like immunoreactivity (cTLI), and canine pancreatic elastase 1 (cE1) after the procedure. ANIMALS: 7 healthy Beagles. PROCEDURES: Clinical examinations were performed and blood samples obtained for serum enzyme determinations before and at intervals (10 minutes; 2, 4, and 6 hours; and 1, 2, and 3 days) after ERP. RESULTS: Repeated clinical examinations revealed no signs of ERP-induced complications in the 7 dogs. Results of repeated laboratory tests indicated a transient increase in serum values of amylase, lipase, and cTLI but not cE1. Mean +/- SD lipase activity increased from 120.7 +/- 116.4 U/L to 423.4 +/- 243.1 U/L at 4 hours after ERP. Median serum cTLI concentration increased from 16.2 microg/L (range, 77 to 26.5 microg/L) to 34.9 microg/L (range, 16.6 to 68.3 microg/L) 10 minutes after ERP. Enzyme values returned to baseline levels at the latest on day 2 in 6 of 7 dogs. Highest values for serum amylase, lipase, and cTLI and their delayed return to baseline values were detected in 1 dog with contrast filling of the pancreatic parenchyma. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that ERP appears to be a safe imaging technique of pancreatic ducts in healthy dogs, although it induced a transient increase in serum values of pancreatic enzymes. In dogs, repeated clinical examinations and serum enzyme determinations can be used to monitor ERP-induced complications such as acute pancreatitis.  相似文献   

11.
Background: The diagnosis of canine pancreatitis is challenging. Clinical presentation often includes nonspecific clinical signs, such as vomiting, anorexia, and abdominal discomfort. Increased serum lipase activity can be indicative of pancreatitis; however, it can also be increased with other conditions. An immunoassay for measurement of canine pancreas‐specific lipase in canine serum that would be suitable for commercial application and provide rapid results would be beneficial. Objective: The goal of this study was to validate the Spec cPL assay, a commercially available ELISA for the quantitative measurement of canine pancreas‐specific lipase. Methods: Dynamic range, dilutional linearity, precision, interfering substances, assay stability, and reproducibility were investigated for analytical validation. The method was compared with the reference assay, canine pancreatic lipase immunoreactivity (cPLI), and included evaluation of a sample population of dogs and bias. Results: Analytical validation showed a dynamic range of 36–954 μg/L; good precision (intra‐ and interassay coefficient of variation <12%); absence of interference from lipid, hemoglobin, or bilirubin; 12‐month kit stability; and good reproducibility. Method comparison showed a positive bias relative to the cPLI reference method; however, the bias can be accommodated by adjustment of decision limits. The upper limit of the reference interval for Spec cPL was determined to be 216 μg/L based on the upper 97.5th percentile of results from 93 clinically healthy, kennel‐housed dogs. Conclusions: Validation data demonstrated that the Spec cPL assay provides reproducible results for canine pancreas‐specific lipase. A readily available assay for measurement of this enzyme allows broader clinical utilization of this analytical tool, generating timely results to aid in the diagnosis of canine pancreatitis.  相似文献   

12.
Leptin is the ob gene product secreted from adipocytes in mammals, and thereby its plasma level reflects body fat content. To establish an assay method for leptin in the dog, rabbit anti-canine leptin antibody was obtained using canine recombinant leptin as an antigen. This antibody reacted to canine leptin much stronger than mouse, rat and human leptins. Sandwich enzyme-linked immunosorbent assay (ELISA) using this antibody was developed. The serum leptin levels of 13 healthy dogs were in a range from 1.4 to 5.6 ng/ml with the mean +/- SEM of 3.0 +/- 0.3 ng/ml.  相似文献   

13.
The objective of this project was to develop and validate a method for concurrent separation and quantification of methylglucose, rhamnose, xylose, sucrose, and lactulose in canine urine by using high pressure anion exchange liquid chromatography and pulsed amperometric detection. The method was validated by evaluating dilutional parallelism, spiking recovery, intra-assay variability, and inter-assay variability. Observed to expected ratios for 3 urine samples, and all sugars, ranged from 77.6% to 106.9% for a 1:2 dilution, 85.2% to 121.4% for a 1:4 dilution, and 91.6% to 163.7% for a 1:8 dilution. Observed to expected ratios for spiking recovery of 3 urine samples, all sugars, and 5 different spiking solutions, ranged from 85.5% to 116.7 % (mean +/- SD, 100.5 +/- 6.0%). The intra-assay coefficients of variation were 1.6%, 3.4%, and 4.7% for methylglucose; 1.6%, 2.0%, and 3.6% for rhamnose; 2.7%, 1.4%, and 1.1% for xylose; 9.8%, 3.4%, and 4.0% for sucrose; and 3.2%, 3.3%, and 3.3% for lactulose. Inter-assay coefficients of variation were 3.2%, 5.7%, and 4.2% for methylglucose; 4.3%, 5.4%, and 6.4% for rhamnose; 3.3%, 5.0%, and 4.2% for xylose; 9.4%, 9.9%, and 9.4% for sucrose; and 6.1%, 4.9%, and 2.7% for lactulose. In conclusion, a method for simultaneous separation and quantification of 5 sugars in canine urine was established and found to be linear, accurate, precise, and reproducible. This method may prove useful in the simultaneous evaluation of gastric permeability, small intestinal permeability, and small intestinal mucosal function in dogs with gastrointestinal disorders.  相似文献   

14.
OBJECTIVE: To develop and analytically validate a radioimmunoassay (RIA) for the quantification of canine calprotectin (cCP) in serum and fecal extracts of dogs. Sample Population-Serum samples (n = 50) and fecal samples (30) were obtained from healthy dogs of various breeds and ages. PROCEDURES: A competitive, liquid-phase, double-antibody RIA was developed and analytically validated by assessing analytic sensitivity, working range, linearity, accuracy, precision, and reproducibility. Reference intervals for serum and fecal cCP concentrations were determined. RESULTS: Sensitivity and upper limit of the working range were 29 and 12,774 microg/L for serum and 2.9 and 1,277.4 microg/g for fecal extracts, respectively. Observed-to-expected ratios for serial dilutions of 6 serum samples and 6 fecal extracts ranged from 95.3% to 138.2% and from 80.9% to 118.1%, respectively. Observed-to-expected ratios for spiking recovery for 6 serum samples and 6 fecal extracts ranged from 84.6% to 121.5% and from 80.3% to 132.1%, respectively. Coefficients of variation for intra-assay and interassay variability were < 3.9% and < 8.7% for 6 serum samples and < 8.5% and < 12.6% for 6 fecal extracts, respectively. Reference intervals were 92 to 1,121 microg of cCP/L for serum and < 2.9 to 137.5 microg of cCP/g for fecal extracts. CONCLUSIONS AND CLINICAL RELEVANCE: The RIA described here was analytically sensitive, linear, accurate, precise, and reproducible for the quantification of cCP in serum and fecal extracts. This assay should facilitate research into the clinical use of serum and fecal cCP measurements in dogs with inflammatory bowel disease.  相似文献   

15.
The British antilewisite butyrate-dithionitrobenzoate (BALB-DTNB) spectrophotometric serum lipase assay was evaluated for precision, accuracy, and diagnostic usefulness in analyzing canine sera. Sera samples from clinically healthy dogs, dogs with experimentally induced pancreatitis, and dogs with spontaneous pancreatitis were analyzed. A titrimetric method of serum lipase determination was used for comparison. Although the BALB-DTNB method was not found to be precise or accurate for determining the lipase activity of canine serum samples, it seemed to be at least as diagnostically useful as the titrimetric procedure. The small sample size requirement and the speed of analysis of the BALB-DTNB procedure are advantages of this method over the titrimetric method, and thus, its use in place of the titrimetric method is justified. A laboratory reference range of 3 to 37 IU/L was determined for canine serum.  相似文献   

16.
Background: Measurement of canine serum insulin has relied on methods developed to measure human insulin. A species‐optimized test for measurement of serum insulin in dogs is now commercially available. Objective: The purpose of this study was to validate the canine ELISA for determination of serum insulin concentration in dogs. Methods: Precision was determined by evaluating intra‐ and interassay coefficient of variation (CV), and accuracy was determined by dilution and spike recovery studies. A method comparison study with samples from 34 clinically healthy dogs and 73 dogs examined for various illnesses and disorders (“patients”) was performed using the canine ELISA and an ELISA for human insulin. Biologic relevance of the canine assay was evaluated by measuring insulin in samples collected from 8 healthy dogs after administration of glucagon. A stability study was preformed with 6 samples stored at 20°C, 4–8°C, and ?20°C. Results: For the canine ELISA, intra‐ and interassay CVs were 4.3–7.8% and 4.4–7.7%, respectively. Mean recovery after dilution was 99% and recovery after spiking with porcine insulin was 116%. The canine and human ELISAs correlated well (r2=.94 for healthy dogs, r2=.88 for patient samples). After glucagon injection serum insulin concentrations increased significantly in 8 dogs. Insulin was stable for 30 days in 6 serum samples stored at ?20°C and in most samples for 8 days at 4–8°C. Insulin was stable for <3 days at room temperature (20°C). Conclusions: The new canine serum insulin ELISA had good precision and accuracy and correlated well with the previously used assay.  相似文献   

17.
S100A12 (calgranulin C) is a Ca(2+)-binding protein that has been proposed to play a central role in both innate and acquired immune responses. In humans, S100A12 has been reported to be increased in serum and/or plasma in patients with various inflammatory disorders, and this protein has been suggested to be a sensitive and specific marker for inflammatory bowel disease (IBD). An immunoassay for S100A12 is currently available for use in humans, but antibodies against the human protein do not cross-react with canine S100A12 (cS100A12). Both sensitive and specific markers for canine patients with systemic or localized inflammatory diseases are currently lacking, thus the aim of this study was to develop and analytically validate a radioimmunoassay (RIA) for the quantification of cS100A12 in serum and fecal specimens and to determine the biological variation of cS100A12 in serum from healthy dogs. A competitive liquid-phase RIA was developed and analytically validated by determining assay working range, dilutional parallelism, spiking recovery, and intra- and inter-assay variability. Reference intervals for serum and fecal concentrations of cS100A12 were established from 124 and 65 healthy dogs, respectively, and components of variation for serum cS100A12 were determined from 11 dogs over 2.6 months. The working range of the assay was 0.6-432.7 μg/L. No cross-reactivity was observed with the cS100A8/A9 protein complex, the closest structural analogues available. Observed-to-expected ratios (O/E) for the serial dilution of serum and fecal extracts ranged from 97.2 to 146.8% and from 75.3 to 129.8%, respectively. O/E for spiking recovery for serum and fecal extracts ranged from 87.8 to 130.4% and from 84.8 to 143.8%, respectively. Coefficients of variation (CV) for intra- and inter-assay variability for sera were ≤ 8.1% and ≤ 7.8%, respectively, and were ≤ 7.8% and ≤ 8.7%, respectively, for fecal extracts. Reference intervals for serum and fecal cS100A12 were 33.2-225.1 μg/L and <24-745 ng/g, respectively. For biological variability testing, analytical, intra-individual, inter-individual, and total CV were 5.7, 29.2, 31.2, and 66.0%, respectively, yielding an index of individuality of 0.95 and a minimum critical difference (p<0.05) for sequential values of 84.9%. The RIA for cS100A12 measurement described here is analytically sensitive and specific, linear, accurate, precise, and reproducible, and will facilitate further research into the clinical utility of quantifying serum and fecal cS100A12 in canine patients with inflammatory diseases. Moderate changes in serum cS100A12 concentrations may be clinically relevant; however, the use of a population-based reference interval may require caution.  相似文献   

18.
The radioimmunoassay (RIA) for trypsin-like immunoreactivity (TLI) is one of the most sensitive and specific tests for detecting exocrine pancreatic insufficiency (EPI). An abnormally low serum TLI concentration (<2.5 ng/ml) indicates end-stage EPI. Although RIA methods can be used to detect canine serum TLI, these procedures are beyond the capabilities of most veterinary clinics and general laboratories. Using monoclonal antibodies (mAbs), we developed an enzyme-linked immunosorbent assay (ELISA) for canine TLI and incorporated it into an immunochromatographic test (ICT) for the diagnosis of EPI. The ELISA was linear over TLI concentrations of 1-100 ng/ml. Levels of intra-assay coefficients of variance (CVs) were 1.8-6.1%, inter-assay CVs were 5.1-9.8%, and the recovery of TLI added to two samples of canine serum ranged from 89 to 111 and 93 to 108%, respectively. Good correlation (correlation coefficient, 0.974) occurred between the TLI values obtained by the ELISA method and those by RIA from 56 clinical samples. Serum TLI values in clinically healthy dogs ranged from 7.8 to 29.2 ng/ml by ELISA, and those from dogs with EPI were 0.0-0.6 ng/ml. The values were 0.0-287.4 ng/ml for dogs with pancreatitis, and those from dogs with gastrointestinal disease were 5.5-58.9 ng/ml. The only statistically significant difference (P<0.01) occurred between the TLI level of healthy dogs and those with EPI. The ICT kit showed high reproducibility, and the TLI values yielding negative results differed significantly (P<0.01) from those returning positive results. The ICT kit yielded negative results (indicating EPI) from clinical serum samples with TLI concentrations of 0.0-4.1 ng/ml by ELISA. Both the ELISA and ICT kit are useful tools in the diagnosis of canine EPI.  相似文献   

19.
BACKGROUND: Increased serum lipase activity has been used historically to support the diagnosis of acute pancreatitis, a common disease in dogs. Most of the lipase assays that are currently in use lack optimum sensitivity and specificity. OBJECTIVE: The objectives of this study were to 1) validate the 1,2-o-dilauryl-rac-glycero-3-glutaric acid-(6'-methylresorufin) ester (DGGR) assay for determination of lipase activity in canine serum and 2) compare results, reference intervals, sensitivity, and specificity of the DGGR assay with a standard 1,2-diglyceride (1,2 DiG) assay for diagnosing acute pancreatitis in dogs. METHODS: Precision, linearity, and interference studies were performed for method validation on a Hitachi 911 analyzer. Lipase results from the DGGR and 1,2 DiG assays were compared by linear regression analysis. Sensitivity, specificity, and diagnostic efficacy were determined for both assays on a population of 30 dogs, 15 of which had acute pancreatitis based on history, clinical signs, and ultrasound findings. RESULTS: Within-run and within-day coefficients of variation (CVs) were low (<3%), with higher day-to-day CVs (< or =14 %). The assay was linear between 8 and 2792 U/L. No significant interference by hemolysis and lipemia was found. Poor correlation was found between the assays (r(s)=0.84). The lipase reference interval was 8-120 U/L for the DGGR assay and 30-699 U/L for the 1,2 DiG assay. Sensitivity and specificity for the diagnosis of pancreatitis were 93% and 53%, respectively, for the DGGR assay and 60% and 73% for the 1,2 DiG assay. Receiver operating characteristic curve analysis showed similar areas under the curve. CONCLUSIONS: On the basis of this study, the DGGR method is considered adequate for assaying serum lipase activity in dogs. The high sensitivity of the DGGR assay suggests it may be a useful screening test for canine pancreatitis.  相似文献   

20.
建立了恩诺沙星残留检测的酶联免疫吸附检测方法(ELISA),并对方法的准确度和灵敏度等指标进行了评价,确定方法的最低检出限(LOD)为1ng/mL,线性范围为1ng/mL-1000ng/mL。以虾肉为样本进行的加标实验表明,在5ng/mL-200ng/mL的加标浓度下,孔间变异系数为9.8%-17.4%,批间变异系数为11.9%-23.4%,添加回收率为60.07%-120.8%。对ELISA和高效液相色谱(HPLC)的检测性能进行了比较,并通过水产品、畜禽等市售食品的同步检测进行了验证,结果表明在实验范围内,二种方法之间呈现较好的相关性,R2=0.9896,这表明建立的ELISA检测方法有较高的可信度是比较高的,可以有效地反映药物的残留情况。  相似文献   

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