BALB-cJ mice immunization with synthetic peptides from LipL32

LOTTERSBERGER, JAVIER; VIARENGO, GASTÓN; GUERRERO, SERGIO ADRIÁN; TARABLA, HÉCTOR; VANASCO, NORMA BIBIANA

Cochin, Querala, India

Congreso; 6th International Leptospirosis Society meeting "LeptoCON 2009"; 2009

International Leptospirosis Society (ILS)

In previous reports, two epitopes for antibody recognition in LipL32 protein from pathogenic Leptospira were identified. Two synthetic peptides including this epitopes were chemically synthesized. The aim of this work was to evaluate specific antibodies induction ability of these two peptides. Peptides P1 (AAKAKPVQKLDDDDDGDDTYKEERHNKYNS) and P2 (YNSLTRIKIPNPPKSFDDLKNIDTKKLVRG) were inoculated in BALB/cJ mice (30-40 g weight) in a stable emulsion (50 ug peptide with Freund adjuvant) by subcutaneous dorsal injection. Six female mice were inoculated (4 times each two weeks) with each peptide, and 3 controls were used in each case. Two weeks after the last injection the mice were decapitated and blood samples obtained according bioethics procedures. Serum sample was titrated in peptide based ELISA test using the corresponding peptide as antigen coated in COSTAR plates with carbonate/bicarbonate buffer (pH 9,6). Serial serum dilution were incubated 1h at 37ºC, washed 5 timers with PBS/Tween 20 and incubated 30 minutes at 37ºC with a 1/4000 dissolution of anti-mouse IgG-peroxidase (Jackson). After 5 washes the reaction was revealed with ready to use TMB (Dako). The results are shows in the table 1. Both peptides shows capacity to induce specific antibodies, but P2 shows more than P1. Individual samples were evaluated in ELISA using different whole-antigen from L. hardjo; L. pomona and L. icterohemorragiae as antigens. Sera from 3 P1 inoculated mice´s and four from P2 reacts with all whole antigens. These results suggest that these two epitopes are relevant in LipL32 antigenicity and the peptides P1 and P2 has a potential useful as a starting point for vaccine design. These results will be extended in challenge to immunizated hamsters.