EVALUATION OF NEUTRALIZING EFFECT OF POLYCLONAL AND MONOCLONAL ANTIBODIES AGAINST Tityus serrulatus VENOM in vitro AND in vivo

ALVARENGA L(1), KALAPOTHAKIS E (2); C. GRANIER (3), DE LIMA ME (1) , OLORTEGUI CC(1), SANTOS RG(4);

(1) Depto de Bioquímica-Imunologia (2) Depto de Biologia Geral- UFMG, Brazil (3) CNRS UMR5160 Centre de Pharmacologie et Biotechnologie pour la Santé Faculté de Pharmacie, Montpellier, France (4) Centro de Desenvolvimento da Tecnologia Nuclear, CDTN/CNEN, Minas Gerais, Brazil

The aim of the present study was to obtain neutralizing monoclonal antibodies (mAbs) against Tityus serrulatus venom by the fusion of SP2/0 murine myeloma cells and spleen cells from BALB/c mice immunized with a toxic fraction (TstFG50) of the Tityus venom conjugated to BSA with glutaraldehyde. A panel of 9 anti-TstFG50 secreting hybridomas was established. The capacity of mAbs to neutralize the TstFG50 toxic fraction toxic was determined by in vivo neutralization assays and by inhibition of the binding of 125I-TsVII to its site on rat brain synaptosomes. In this work we used an in vivo neutralization assay based on the envenomation with a radiolabeled version of the main toxic component of the venom, 125I-TsVII, after pre-injection of monoclonal antibodies. 125I-TsVII specific binding on synaptosomes was strongly inhibited by several monoclonal antibodies. In vivo biodistribution in mice showed that the radiolabeled TsVII accumulated in lung and heart tissue, cleared rapidly from the blood, and was excreted significantly via the kidneys. Monoclonal antibodies MAbTs1, which react with peptide 26 of TsIV (KKSKDKKADSGYSYW), peptide 30 of TsVII (KKGSSGYSAWPASYS) and peptide 31 of TsNTxP (KKGSSGYSAWPASYS), was able to inhibit 35% of the specific interaction of 125I-TsVII with synaptosomes and neutralize the specific uptake, in vivo, in target organs responsible for the envenomation symptoms. In this work we produced monoclonal antibodies capable to inhibit the specific interaction of the toxic protein (TsVII) with sodium channels present on brain synaptosomes and neutralize its in vivo uptake by target organs involved in the scorpion envenomation symptoms.

KEY WORDS: Tytius venom, monoclonal antibodies, neutralization

FINANCIAL SUPPORT: CNPq, CDTN/CNEN

CORRESPONDENCE TO: Raquel Gouvêa Dos Santos, Lab. Radiobiologia; CDTN/CNEN; C. P. 941; 30123-970; Belo Horizonte - MG – Brasil. santosr@cdtn.br