Authors : Arcelia Alvarado Islas , Angel Horacio Sandoval Trujillo , Victor R. Tenorio Gutierrez , Rogelio Alonso Morales , Octavio de Paz Villafan and Alvaro Aguilar
Abstract: The AL27 crotalic fraction is a lectin recognized by cell receptors and then introduced into the cytoplasm. In the present research a new transfection promoter agent for a non viral expression vector was used, using as a model a plasmid expressing the gene that codifies for the G glycoprotein of the rabies virus. The objective was to evaluate the ability of AL27 to increase the rabies plasmid transfection in in vitro and in vivo conditions. The AL27 fraction was isolated from the venom of Agkistrodon piscivorus by HPLC-RP. AL27 maintained its recognition by sialic acid receptors up to a titer of 64 HU. The pC38 plasmid was selected from a panel of 18 clones constructed by inserting the gene that codifies for glycoprotein G of rabies strain HQIMSS99 into the commercial pCI-neo expression vector. In vitro results showed that pC38, with or without AL27, transfected CHO cells under the calcium-phosphate system and using Confocal Microscopy (CM) increased fluorescence (+++) was observed with the mixture pC38/AL27, than with pC38 alone (++). Flow cytometry (FC) revealed expression percentages of 27.78 and 38.66% for pC38 and pC38/AL27, respectively. The in vivo experiment was performed using three groups of BALB/C mice: group 1 was inoculated with pC38 in the tibial muscle; group 2 with pC38 and AL27 and group 3, negative control, with only PBS. Tissues showed the highest expression by CM (+++) when using pC38/AL27. Antibodies detected ranged from 0.75-1.2 international units (IU) mL 1 for pC38 and from 1.1-1.27 IU mL 1 for pC38/AL27. Results showed that AL27 promoted cell entrance mechanisms, generating a better expression level of the gene that codifies for the G glycoprotein of the rabies virus and consequently enhancing the stimulation of the immune response.
Arcelia Alvarado Islas , Angel Horacio Sandoval Trujillo , Victor R. Tenorio Gutierrez , Rogelio Alonso Morales , Octavio de Paz Villafan and Alvaro Aguilar , 2008. Crotalic Venom Fraction as Promoter of the Transfection Mechanism of a Genic Vaccine (Naked DNA) Against Rabies. Journal of Animal and Veterinary Advances, 7: 1045-1055.