Publication: Hepatit B yüzey antijenini taklit eden anti-idiyotipik monoklonal antikorların üretilmesi ve bağışıklama yeteneklerinin In-Vivo sistemde incelenmesi
Abstract
Hepatit B virüsü (HBV) akut ve kronik hepatit B, karaciğer sirozu ve hepatoselüler karsinoma gibi hastalıkların en önemli etkenidir. HBV ile enfekte olan bir kişi için günümüzde etkin bir tedavi söz konusu değildir. HBV enfeksiyonuna karşı, toplum sağlığının korunmasında en etkin yöntem aşılamadır. Mevcut Hepatit B aşıları, bağışık cevabın tam oluşması için 3 doz uygulanmasına rağmen yetişkinlerin yaklaşık %5'inde cevap oluşturamamaktadır. Bu sebeplerle geliştirilmeye başlanan yeni aşı yaklaşımlarından biri etken antijenini taklit eden anti-idiyotipik antikorların kullanılmasıdır. Tez çalışmasında, Hepatit B virüsü yüzey antijenine (HBsAg) karşı geliştirilen anti-HBsAg (aHBsAg) monoklonal antikorları kullanarak hibridoma teknolojisi ile anti-idiyotipik aHBsAg (aid-aHBsAg) monoklonal antikorlarının üretilmesi ve bu aid-aHBsAg monoklonal antikorlarının HBsAg'ye karşı antikor yanıtı (aHBsAg) oluşturma yeteneklerinin in-vivo sistemde gösterilmesi amaçlandı. Bunun için TÜBİTAK-GMBAE Hibridoma Laboratuarında HBsAg'ye karşı geliştirilmiş IgG izotipindeki MAM-A1, 2B7 ve 9D12 monoklonal antikorları (aHBsAg) kullanılarak fareler bağışıklandı. En yüksek bağışık cevabı gösteren fareler sırasıyla füzyona alındı. Füzyon sonrası, IgG2b izotipinde 1F11; IgG3 izotipinde 11E2; IgM izotipinde ise 4F11 ve 5B11 aid-aHBsAg monoklonal antikorları elde edildi. IgG tipi antikorlar Protein-G immün-afinite, IgM tipi antikorlar ise S-300 jel filtrasyon kolon kromatografisi ile saflaştırılarak farelere çeşitli dozlarda enjekte edildi. Enjeksiyon sonrası, 11E2 ve 4F11 aid-aHBsAg monoklonal antikorlarının ticari aşıya benzer aHBsAg cevabı oluşturduğu gözlendi. Elde edilen verilere göre aid-aHBsAg monoklonal antikorlarının, aHBsAg antikor cevabı oluşturmada etkili olabileceği ve tanımlanan yöntemlerin alternatif aşı üretiminde kullanılabileceği sonucuna varıldı. Monoklonal antikor; Hibridoma teknolojisi; Anti-idiyotip; Hepatit B; Bağışık cevap Production of anti-idiotypic monoclonal antibodies that mimic the hepatitis B surface antigen and investigation of their immunogenic capability in-vivo
Hepatitis B virus (HBV) is the major cause of acute and chronic hepatitis B, liver cirrhosis, and hepatocellular carcinoma. Today, there is no effective treatment for people infected by HBV. The most potent method for protecting society against HBV infection is vaccination. Although as much as 3 dose applications of the current hepatitis B vaccine are performed to obtain the whole immune response, about %5 of adults are non-responders. Therefore, one of the new approaches for developing vaccines is utilizing anti-idiotypic antibodies, which mimic the antigen. In this study, we aimed to develop anti-idiotypic anti-HBV surface antigen (aid-aHBsAg) monoclonal antibodies by employing anti-HBsAg (aHBsAg) monoclonal antibodies which had previously been produced against HBV surface antigen (HBsAg) by means of hybridoma technology and to demonstrate the capability of these aid-aHBsAg monoclonal antibodies for developing an immune response against HBsAg in-vivo. To that end, MAM-A1, 2B7 and 9D12 monoclonal antibodies of IgG isotype that had been produced against HBsAg in TÜBİTAK-GMBAE Hybridoma Laboratory were used for immunizing mice. The most stimulated mice were selected for the fusion. Following the fusion, four aid-aHBsAg monoclonal antibodies which, namely 1F11 of IgG2b isotype, 11E2 of IgG3 isotype, and 4F11 and 5B11 of IgM isotype, were produced. Antibodies of IgG isotype were purified by Protein-G immuno-affinity while antibodies of IgM isotype were purified by S-300 gel filtration column chromatography. Subsequently, different doses of these antibodies were injected into mice. Postinjection, it was demonstrated that 11E2 and 4F11 aid-aHBsAg monoclonal antibodies induced an aHBsAg response in mice similar to that of commercial vaccine. These results show that aid-aHBsAg monoclonal antibodies could be effective in inducing antibody response against HBsAg and techniques that were explained could be used as an alternative way of vaccine production. Monoclonal antibody; Hybridoma technology; Anti-idiotype; Hepatitis B; Immune response
Hepatitis B virus (HBV) is the major cause of acute and chronic hepatitis B, liver cirrhosis, and hepatocellular carcinoma. Today, there is no effective treatment for people infected by HBV. The most potent method for protecting society against HBV infection is vaccination. Although as much as 3 dose applications of the current hepatitis B vaccine are performed to obtain the whole immune response, about %5 of adults are non-responders. Therefore, one of the new approaches for developing vaccines is utilizing anti-idiotypic antibodies, which mimic the antigen. In this study, we aimed to develop anti-idiotypic anti-HBV surface antigen (aid-aHBsAg) monoclonal antibodies by employing anti-HBsAg (aHBsAg) monoclonal antibodies which had previously been produced against HBV surface antigen (HBsAg) by means of hybridoma technology and to demonstrate the capability of these aid-aHBsAg monoclonal antibodies for developing an immune response against HBsAg in-vivo. To that end, MAM-A1, 2B7 and 9D12 monoclonal antibodies of IgG isotype that had been produced against HBsAg in TÜBİTAK-GMBAE Hybridoma Laboratory were used for immunizing mice. The most stimulated mice were selected for the fusion. Following the fusion, four aid-aHBsAg monoclonal antibodies which, namely 1F11 of IgG2b isotype, 11E2 of IgG3 isotype, and 4F11 and 5B11 of IgM isotype, were produced. Antibodies of IgG isotype were purified by Protein-G immuno-affinity while antibodies of IgM isotype were purified by S-300 gel filtration column chromatography. Subsequently, different doses of these antibodies were injected into mice. Postinjection, it was demonstrated that 11E2 and 4F11 aid-aHBsAg monoclonal antibodies induced an aHBsAg response in mice similar to that of commercial vaccine. These results show that aid-aHBsAg monoclonal antibodies could be effective in inducing antibody response against HBsAg and techniques that were explained could be used as an alternative way of vaccine production. Monoclonal antibody; Hybridoma technology; Anti-idiotype; Hepatitis B; Immune response