Development and evaluation of a gold nanoparticle based Lateral Flow assay (LFA) strip test for detection of Brucella spp.

The widely used serodiagnostic test (RBPT, CFT , I-ELISA and FPA) for diagnosis of brucellosis cannot detect vertically infected or carrier animals that are seronegative, a persistent source of infection to other susceptible animals in the herd. For reducing transmission of disease within the herd, these animals must be detected using a rapid, sensitive, user friendly penside diagnostic test. In the present study, Lateral Flow immunoassay (LFA) strip test was developed for detection of Brucella spp. from clinical samples (bovine aborted foetal stomach contents). The LFA strip was fabricated by printing anti-Brucella polyclonal antibodies (PAb) and anti-bovine antibodies IgG on test and control line, respectively. For conjugation, colloidal gold nanoparticles (30 nm GNP, Sigma, USA) were conjugated with anti-brucella PAb. The LFA strip test was able to detect 10 7 cfu/ml B.abortus S99 inactivated organism in PBS and it did not exhibit any cross reactivity with selected non Brucella pathogens. To further validate, 115 clinical specimens were tested using LFA strip test. The relative sensitivity (DSn) and relative specificity (DSp) of LFA strip test was determined by ROC curve analysis using PCR and culture method as reference standard. DSn and DSp of LFA strip test was observed as 78.57% (95%CI: 49.2–95.3); 93.07% (95%CI: 86.2–97.2) and 80.0% (95%CI:51.9–95.7); 94.0% (95%CI:0.795–0.925) using culture and PCR as reference diagnostic tests, respectively. It may be concluded that, the LFA strip test can be used as a rapid penside diagnostic test for screening of brucellosis. To the best of our knowledge, this is the first report on development of GNP based LFA strip test for detection of Brucella spp. from bovine aborted fetal content samples. • Seronegative brucellosis infected animals play a crucial role in perpetuating the disease in the herd. • This novel LFA strip test could detect as low as 10 7 cfu/ml B.abortus S99 inactivated organism in PBS. • This test did not exhibit any cross reactivity with non-Brucella pathogens, indicated good sensitivity and specificity. • The LFA strip test is simple, easy to perform and results can be interpreted with naked eyes within 15-20 min.