|680-3||Recombinant peptides that mimic the Chlamydia trachomatis selected by phage display as novel epitope-based diagnostic probes|
|Autores:||Luiz Fernando Almeida Machado (UFPA - Universidade Federal do Pará) ; Fabiana de Almeida Araújo Santos (UFU - Universidade Federal de Uberlândia) ; Mayara Ingrid Sousa Lima (UFU - Universidade Federal de Uberlândia) ; Vivian Alonso Goulart (UFU - Universidade Federal de Uberlândia) ; Sebastião Marcos Tafuri (UFU - Universidade Federal de Uberlândia) ; Marluisa Oliveira Guimarães Ishak (UFPA - Universidade Federal do Pará) ; Vania Nakauth Azevedo (UFPA - Universidade Federal do Pará) ; Antonio Carlos Rosário Vallinoto (UFPA - Universidade Federal do Pará) ; Ricardo Ishak (UFPA - Universidade Federal do Pará) ; Carlos Ueira Vieira (UFU - Universidade Federal de Uberlândia) ; Luiz Ricardo Goulart Filho (UFU - Universidade Federal de Uberlândia) |
The development of new diagnostic assays for detection of specific antibodies to microbial pathogens is necessary and the phage display has been shown a powerful methodology to find new targets to be used in immunoassays. Chlamydia trachomatis causes a spectrum of diseases in humans, including trachoma (a leading cause of blindness) and sexually transmitted infections that often result in severe sequelae in women such as infertility, ectopic pregnancy and chronic pelvic pain. With the phage display method, previously unknown conformational and linear epitopes of high affinity monoclonal and polyclonal antibodies have been successfully identified. In this study, IgG from patients infected with C. trachomatis and negative controls were covalently coupled to protein G-magnetic beads. After coupling, beads were washed and incubated with a PhD-12 library of random hepta-peptides expressed at the amino terminal of pIII coat protein of the filamentous bacteriophage M13, which has a diversity of 109 peptides ( New England BioLabs, Inc) for biopanning. Our goal was to obtain specific peptide ligands to IgG that mimic C. trachomatis epitopes. After selection, 92 clones were tested in ELISA assays against sera from C. trachomatis infected patients and negative controls, and the highly reactive clones had their DNA sequenced and translated. Additional immunoassays were performed for those clones with valid sequences, which showed significant differences between patients and controls. We have further performed an in silico analysis and found relevant linear and structural alignments to viral bacterial. The direct use of selected phage-fused mimotopes present a significant improvement in C. trachomatis diagnosis, and may become readily use for screening purposes in many different immunoassays.
Palavras-chave: Chlamydia trachomatis, Mimotopes, Phage Display