Baker Institute for Animal Health, College of Veterinary Medicine, Cornell University, thaca, NY, 14853, USA.
Department of Clinical Sciences, College of Veterinary Medicine, Cornell University, Ithaca, NY, 14853, USA.
Sci Rep. 2017 Oct 5;7(1):12713. doi: 10.1038/s41598-017-12725-5.
In response to immunization, B-cells generate a repertoire of antigen-specific antibodies. Antibody-based immunotherapies hold great promise for treating a variety of diseases in humans. Application of antibody-based immunotherapy in cats is limited by the lack of species-specific complete sequences for mRNAs encoding rearranged heavy and light chain immunoglobulins in B cells. To address this barrier, we isolated mRNAs from feline peripheral blood mononuclear cells (PBMCs), and used available immunoglobulin sequences and 5' and 3' RACE to clone and sequence heavy and light chain immunoglobulin mRNAs. We recovered mRNA from PBMCs from two cats, cloned and sequenced the variable and constant domains of the feline heavy chains of IgG1a (IGHG1a), IgG2 (IGHG2), and IgA (IGHA), and the light chains (lambda and kappa). Using these sequences, we prepared two bicistronic vectors for mammalian expression of a representative feline heavy (IGHG1a) together with a light (lambda or kappa) chain. Here we report novel feline Ig sequences, a technique to express antigen-specific felinized monoclonal antibodies, and the initial characterization of a functional felinized monoclonal antibody against feline panleukopenia virus.
针对免疫反应,B 细胞会产生一系列抗原特异性抗体。基于抗体的免疫疗法在治疗人类多种疾病方面具有巨大的潜力。由于缺乏猫科动物 B 细胞中重链和轻链免疫球蛋白基因重排的特异性完整 mRNA 序列,基于抗体的免疫疗法在猫科动物中的应用受到限制。为了解决这一障碍,我们从猫外周血单核细胞 (PBMC) 中分离出 mRNA,并利用现有的免疫球蛋白序列和 5' 和 3' RACE 克隆和测序重链和轻链免疫球蛋白 mRNA。我们从两只猫的 PBMC 中回收 mRNA,克隆并测序了 IgG1a(IGHG1a)、IgG2(IGHG2)和 IgA(IGHA)的重链以及轻链(lambda 和 kappa)的可变和恒定区。使用这些序列,我们为哺乳动物表达准备了两个双顺反子载体,用于表达代表性的猫科动物重链(IGHG1a)和轻链(lambda 或 kappa)。在这里,我们报告了新型的猫科动物 Ig 序列、一种表达抗原特异性猫科动物化单克隆抗体的技术,以及针对猫泛白细胞减少症病毒的功能性猫科动物化单克隆抗体的初步表征。