Testing different antigen capture ELISA formats for detection of Leptospira spp. in human blood serum.
Leptospirosis is an infectious disease caused by pathogenic spirochetes of the genus Leptospira. The illness is characterized by an acute bacteremic phase followed by an immune phase, in which specific antibodies are found in blood and leptospires are eliminated in urine. Novel diagnostic strategies for use in the acute phase of leptospirosis are needed since clinical manifestations in this phase mimic other feverish tropical diseases. In the present study, mAbs and polyclonal IgY were used in the standardization of three different antigen capture ELISA formats for direct detection of leptospires in human blood during the acute phase of the disease. Detection limit of leptospires in experimentally contaminated human sera ranged from 10(5) to 10(7) cells ml(-1) in the different formats. The ELISA format with the best performance was able to detect 10(5) leptospires ml(-1) in human sera using a mAb against LipL32, the major outer membrane protein of pathogenic leptospires, as capture antibody, and a biotinylated polyclonal IgY against a pathogenic serovar of L. interrogans Icterohamorrhagiae as detection antibody. By increasing the degree of IgY biotinylation this detection limit could be improved to make the assay clinically useful.
Vasconcellos
Flávia A
FA
Centro de Biotecnologia, Universidade Federal de Pelotas, Cx. Postal 354, 96010-900, Pelotas, RS, Brazil.
Coutinho
Mariana L
ML
da Silva
Everton F
EF
Fernandes
Cláudia P H
CP
Monte
Leonardo G
LG
Seyffert
Núbia
N
Dellagostin
Odir Antônio
OA
Aleixo
José Antonio G
JA
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D000942:Antigens, Bacterial / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D007919:Leptospira / Q000276:immunology* / Q000302:isolation & purification; D007922:Leptospirosis / Q000175:diagnosis* / Q000276:immunology; D057230:Limit of Detection; D012680:Sensitivity and Specificity
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial