Production of Human Antibody Fragments Binding to Melittin and Phospholipase A2 in Africanised Bee Venom: Minimising Venom Toxicity

The hybrid created from the crossbreeding of European and African bees, known as the Africanised bee, has provided numerous advantages for current beekeeping. However, this new species exhibits undesirable behaviours, such as colony defence instinct and a propensity to attack en masse, which can result in serious accidents. To date, there is no effective treatment for cases of Africanised bee envenomation. One promising technique for developing an efficient antivenom is the use of phage display technology, which enables the production of human antibodies, thus avoiding the complications of serum therapy, such as anaphylaxis and serum sickness. The aim of this study was to produce human monoclonal single-chain Fv (scFv) antibody fragments capable of inhibiting the toxic effects of Africanised bee venom. We conducted four rounds of selection of antibodies against the venom and three rounds of selection of antibodies against purified melittin. Three clones were selected and tested by enzyme-linked immunosorbent assay to verify their specificity for melittin and phospholipase A2. Two clones (C5 and C12) were specific for melittin, and one (A7) was specific for phospholipase A2. In a kinetic haemolytic assay, these clones were evaluated individually and in pairs. The A7-C12 combination had the best synergistic effect and was chosen to be used in the assays of myotoxicity inhibition and lethality. The A7-C12 combination inhibited the in vivo myotoxic effect of the venom and increased the survival of treated animals.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo/Instituto Nacional de Ciencia e Tecnologia em Toxinas (FAPESP/INCTTOX, Sao Paulo Research Foundation/National institute of Science and Technology in Toxicology)Fundacao de Amparo a Pesquisa do Estado de Sao Paulo/Instituto Nacional de Ciencia e Tecnologia em Toxinas (FAPESP/INCTTOX, Sao Paulo Research Foundation/National institute of Science and Technology in Toxicology) [573790/2008-6]Fundacao de Apoio ao Ensino, Pesquisa e Assistencia (FAEPA, Foundation for the Support of Instruction, Research and Treatment)Fundacao de Apoio ao Ensino, Pesquisa e Assistencia (FAEPA, Foundation for the Support of Instruction, Research and Treatment)Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq, Brazilian National Council for Scientific and Technological Development)Brazilian Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq, National Council for Scientific and Technological Development