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Evaluation of a monoclonal antibody–based dot-blot ELISA for detection of Leptospira spp in bovine urine samples

Junpen SuwimonteerabutrDepartment of Microbiology and Immunology, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand 10400.

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Wanpen ChaicumpaDepartment of Medical Technology, Faculty of Allied Health Sciences, Thammasat University, Rangsit Center, Pathum-thani, Thailand 12121.

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Patcharin SaengjarukDepartment of Pathology, Faculty of Medicine, Srinakharinwirot University, Bangkok, Thailand 10110.

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Pramuan TapchaisriDepartment of Medical Technology, Faculty of Allied Health Sciences, Thammasat University, Rangsit Center, Pathum-thani, Thailand 12121.

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Manas Chongsa-nguanDepartment of Microbiology and Immunology, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand 10400.

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Thareerat KalambahetiDepartment of Microbiology and Immunology, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand 10400.

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Pongrama RamasootaDepartment of Social Medicine and Environment, Faculty of Tropical Medicine, Mahidol University, Bangkok, Thailand 10400.

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Yuwaporn SakolvareeDepartment of Medical Technology, Faculty of Allied Health Sciences, Thammasat University, Rangsit Center, Pathum-thani, Thailand 12121.

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Prachin VirakulDepartment of Obstetrics and Gynecology, Faculty of Veterinary Medicine, Chulalongkorn University, Bangkok, Thailand 10300.

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Abstract

Objective—To evaluate the efficacy of a novel monoclonal antibody (MAb)–based dot-blot ELISA for detection of Leptospira antigens in urine samples of cattle.

Sample Population—Blood and urine samples of 45 test cattle from 5 farms in Chonburi province and 20 control cattle from 2 farms in Khon Kaen province in Thailand.

Procedure—Blood and urine samples were assayed (microscopic agglutination test and urine antigen test) for Leptospira infection by use of an MAb–based dotblot ELISA, and results for the ELISA were compared with those for dark-field microscopy (DFM), microbial culture, and a polymerase chain reaction (PCR) assay.

Results—All urine samples with positive results for DFM, microbial culture, PCR assay, or > 1 of these tests also had positive results when tested by use of the MAb–based dot-blot ELISA, except for 1 sample that had positive results only for the PCR assay. Detection limits of the dot-blot ELISA were 103 leptospires/ mL of urine and 9.3 ng of Leptospira homogenate. Comparison revealed that the diagnostic sensitivity, specificity, efficacy (accuracy), positive predictive value, and negative predictive value for the ELISA were in agreement with results for DFM (100%, 72.72%, 80%, 57.14%, and 100%, respectively), microbial culture (100%, 61.54%, 66.62%, 28.57%, and 100%, respectively), and PCR assay (95.45%, 100%, 91.77%, 100%, and 95.83%, respectively).

Conclusions and Clinical Relevance—The MAb–based dot-blot ELISA is suitable as a tool for detecting leptospires in urine samples of cattle. (Am J Vet Res 2005;66:762–766)

Abstract

Objective—To evaluate the efficacy of a novel monoclonal antibody (MAb)–based dot-blot ELISA for detection of Leptospira antigens in urine samples of cattle.

Sample Population—Blood and urine samples of 45 test cattle from 5 farms in Chonburi province and 20 control cattle from 2 farms in Khon Kaen province in Thailand.

Procedure—Blood and urine samples were assayed (microscopic agglutination test and urine antigen test) for Leptospira infection by use of an MAb–based dotblot ELISA, and results for the ELISA were compared with those for dark-field microscopy (DFM), microbial culture, and a polymerase chain reaction (PCR) assay.

Results—All urine samples with positive results for DFM, microbial culture, PCR assay, or > 1 of these tests also had positive results when tested by use of the MAb–based dot-blot ELISA, except for 1 sample that had positive results only for the PCR assay. Detection limits of the dot-blot ELISA were 103 leptospires/ mL of urine and 9.3 ng of Leptospira homogenate. Comparison revealed that the diagnostic sensitivity, specificity, efficacy (accuracy), positive predictive value, and negative predictive value for the ELISA were in agreement with results for DFM (100%, 72.72%, 80%, 57.14%, and 100%, respectively), microbial culture (100%, 61.54%, 66.62%, 28.57%, and 100%, respectively), and PCR assay (95.45%, 100%, 91.77%, 100%, and 95.83%, respectively).

Conclusions and Clinical Relevance—The MAb–based dot-blot ELISA is suitable as a tool for detecting leptospires in urine samples of cattle. (Am J Vet Res 2005;66:762–766)