by Western blotting was performed as previously mentioned. anti p430 exhibited weak viral neutralization capacity on the same samples (31.25%). Furthermore Ab mixes containing anti p430 exhibited reduced viral neutralization properties. From these experiments one could predict that neutralization by Abs towards different E2-epitopes varies considerably and success in the enrichment of neutralization epitope-specific antibodies may be accompanied by favorable results in combating HCV infection. Also, E2 conserved peptides p517 and p412 represent potential components of a candidate peptide vaccine against HCV infection. Keywords: Hepatitis C virus (HCV), anti E2 antibodies, neutralizing antibodies, In vitro culture model for HCV, candidate peptide vaccine for HCV Introduction Hepatitis C Virus (HCV) is a global health problem that affects almost 3% of the world’s population [1] and not less than 15% of the Egyptian population [2]. Individuals with chronic HCV infection usually remain asymptomatic and undiagnosed for decades before chronic hepatitis leads to severe fibrosis and cirrhosis, hepatic failure, or hepatocellular carcinoma [3-7]. These long-term complications made HCV one of the leading emerging infectious diseases worldwide. The current antiviral regimen, a combination of pegylated interferon and ribavirin, is curative in about half of treated patients depending on the viral and/or host factors. Additionally, this regimen is expensive, requires prolonged therapy, sometimes with serious side effects and A 438079 hydrochloride only a fraction of those with chronic HCV infections meet the criteria for treatment [8]. Viral proteins are recognized as nonself by the host’s immune system and induce the production Rabbit polyclonal to PDE3A of antibodies. During the natural course of infection, a large number of antibodies are produced. The vast majority of antibodies induced have no antiviral activity, either because they are elicited by degraded or incompletely processed proteins released from dying cells or because they are directed against epitopes that do not play any role in the virus entry process “non-neutralizing antibodies”. A small proportion of antibodies termed “neutralizing antibodies” are able A 438079 hydrochloride to target exposed epitopes of the viral structural proteins and neutralize the infectious virus by preventing or controlling viral infection [9,10]. During the chronic phase of HCV infection, most HCV-infected patients develop high-titer of antibodies. Paradoxically, these antibodies were not able to control HCV infection which may be attributed to the generation of non-neutralizing HCV-specific antibodies that compete with neutralizing Abs and reduce their effectiveness. Such antibodies have been reported in other viral infections in which highly immunogenic non-neutralizing epitopes mislead the humoral immune response contributing to viral escape from neutralization [11]. Several observations support the hypothesis that neutralizing antibodies may help control HCV replication [12,13]. Synthetic peptide based vaccines were shown to generate specific Abs capable of neutralizing HCV infections [14,15]. In the present study, we utilized large level multiple sequence positioning of E2 to design genetically A 438079 hydrochloride conserved peptides from viral envelop proteins (particularly among type 4 isolates predominant in Egypt). The aim of this work is to develop monospecific polyclonal Abdominal muscles in caprines against the 3 chosen conserved peptides derived from E2 glycoprotein and to test the immunogenic and viral neutralizing properties of each Ab using assays depending on obstructing of viral infectivity to hepatoma cell collection. Based on the acquired results, p412 and p517 represent candidate peptides for further assessment as potential restorative/prophylactic immunogens. Materials and methods Authorization ethics This study was authorized by the Review Table of National Study Center, Egypt Design and synthesis of HCV E2 conserved peptides Three peptides were designed and synthesized as previously explained [16]. Peptides were all derived from the C-terminal region of HVR-1 and designated p412 [a.a 412-419], p430 [a.a 430-547] and p517 [a.a 517-531]. Immunization of caprines, production and purification of polyclonal antibodies Six goats were immunized with either of the synthetic peptides p412, p 430 or p517 (2 animals for each peptide). Two goats were injected with 2 ml saline remedy at the same time intervals of immunization protocol to serve as settings. The immunizing doses/goat were 1.5 mg of the peptide. Each linear peptide was emulsified with equivalent volume of Freund’s total adjuvant and was injected subcutaneously in three different sites within the same animal. On days 15 and 28, blood samples were collected and then each goat was reimmunized with a second booster dose of the same peptide emulsified with equivalent volume of Freund’s incomplete adjuvant. The goats were sacrificed after 96 days from your last injection and A 438079 hydrochloride the blood was collected to quantify the titer.
