title: Visual detection of canine parvovirus based on loop-mediated isothermal amplification combined with enzyme-linked immunosorbent assay and with lateral flow dipstick.
abstract: Loop-mediated isothermal amplification (LAMP) combined with enzyme-linked immunosorbent assay (LAMP-ELISA) and with lateral flow dipstick (LAMP-LFD) are rapid, sensitive and specific methods for the visual detection of clinical pathogens. In this study, LAMP-ELISA and LAMP-LFD were developed for the visual detection of canine parvovirus (CPV). For LAMP, a set of four primers (biotin-labeled forward inner primers) was designed to specifically amplify a region of the VP2 gene of CPV. The optimum time and temperature for LAMP were 60 min and 65°C, respectively. The specific capture oligonucleotide probes, biotin-labeled CPV probe for LAMP-ELISA and fluorescein isothiocyanate-labeled CPV probe for LAMP-LFD were also designed for hybridization with LAMP amplicons on streptavidin-coated wells and LFD strips, respectively. For the comparison of detection sensitivity, conventional PCR and LAMP for CPV detection were also performed. The CPV detection limits by PCR, PCR-ELISA, LAMP, LAMP-ELISA and LAMP-LFD were 10(2), 10(2), 10(-1), 10(-1) and 10(-1) TCID50/ml, respectively. In tests using artificially contaminated dog fecal samples, the samples with CPV inoculation levels of ≥1 TCID50/ml gave positive results by both LAMP-ELISA and LAMP-LFD. Our data indicated that both LAMP-ELISA and LAMP-LFD are promising as rapid, sensitive and specific methods for an efficient diagnosis of CPV infection.
authors: Animal Technology Laboratories, Agricultural Technology Research Institute, Hsinchu City, Taiwan, R.O.C.
authors: Yu-Ling
authors: YL
authors: Sun
authors: Chon-Ho
authors: CH
authors: Yen
authors: Ching-Fu
authors: CF
authors: Tu
mesh_terms: D000818:Animals; D001483:Base Sequence; D036022:Capsid Proteins / Q000235:genetics; D017931:DNA Primers / Q000235:genetics; D004283:Dog Diseases / Q000175:diagnosis* / Q000821:virology*; D004285:Dogs; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods / Q000662:veterinary*; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques / Q000379:methods / Q000662:veterinary*; D015345:Oligonucleotide Probes; D010322:Parvoviridae Infections / Q000175:diagnosis / Q000662:veterinary*; D017993:Parvovirus, Canine / Q000302:isolation & purification*; D012680:Sensitivity and Specificity; D016415:Sequence Alignment
chemical_list: D036022:Capsid Proteins; D017931:DNA Primers; D015345:Oligonucleotide Probes