HYBRIDOMA TECHNOLOGY FOR PRODUCTION OF MONOCLONAL ANTIBODIES
HYBRIDOMA TECHNOLOGY FOR PRODUCTION OF MONOCLONAL ANTIBODIES
复制标题
用于生产单克隆抗体的杂交瘤技术
DOI：
--
复制
发表时间：
2010
期刊：
影响因子：
--
通讯作者：
Shivanand Pandey
中科院分区：
文献类型：
--
作者：
Shivanand Pandey
杂交瘤是已经被工程化以大量产生所需抗体的细胞。为了生产单克隆抗体，从已经用相关抗原攻击的动物的脾脏中取出B细胞。然后将这些B细胞与骨髓瘤肿瘤细胞融合，这些肿瘤细胞可以在培养物中无限生长（骨髓瘤是一种B细胞癌症）。这种融合是通过使细胞膜更具渗透性来进行的。融合的杂交细胞（称为杂交瘤），是癌细胞，将迅速和无限繁殖，并将产生大量的所需抗体。它们必须被选择，随后通过有限稀释进行克隆。单克隆抗体的生产首先由塞萨尔米尔斯坦，乔治斯J. F. 1975年，科勒和尼尔斯·卡伊·杰恩。通过在选择性培养基，特别是含有1x浓度HAT的培养基中培养新融合的原代杂交瘤细胞约10-14天来进行选择。使用HAT后，通常需要使用含HT的培养基。在鉴定阳性原代杂交瘤细胞后进行克隆。有限稀释克隆。虽然有些人可能认为IL-6对这一步骤至关重要，但没有必要添加昂贵的补充剂，而是在第一周使用50%热灭活的FBS。单菌落威尔斯孔筛选后，向克隆培养板中加入10% FBS DMEM。
Hybridomas are cells that have been engineered to produce a desired antibody in large amounts. To produce monoclonal antibodies, Bcells are removed from the spleen of an animal that has been challenged with the relevant antigen. These B-cells are then fused with myeloma tumor cells that can grow indefinitely in culture (myeloma is a B-cell cancer). This fusion is performed by making the cell membranes more permeable. The fused hybrid cells (called hybridomas), being cancer cells, will multiply rapidly and indefinitely and will produce large amounts of the desired antibodies. They have to be selected and subsequently cloned by limiting dilution. Supplemental media containing Interleukin-6 (such as briclone) are essential for this step.The production of monoclonal anti-bodies was first invented by Cesar Milstein, Georges J. F. Kohler and Niels Kaj Jerne in 1975. Selection occurs via culturing the newly fused primary hybridoma cells in selective-media, specifically media containing 1x concentration HAT for roughly 10–14 days. After using HAT it is often desirable to use HT containing media. Cloning occurs after identification of positive primary hybridoma cells. Clone by limited dilution. While some may believe that IL-6 is essential for this step, it is not necessary to add that expensive supplement, rather use 50% heat-inactivated FBS for the first week. Add 10% FBS DMEM to the clone culture plate after screening for single colony wells.