A Rapid Method for Antigen-Specific Hybridoma Clone Isolation,Analytical Chemistry

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A Rapid Method for Antigen-Specific Hybridoma Clone Isolation
Analytical Chemistry ( IF 6.7 ) Pub Date : 2018-01-17 00:00:00 , DOI: 10.1021/acs.analchem.7b04595
Xiuqing Li ₁ , Hongfen Bian ₁ , Siming Yu ₂ , Wei Xiao ₁ , Jianying Shen ₃ , Caifeng Lan ₁ , Kenan Zhou ₁ , Caihong Huang ₁ , Lei Wang ₁ , Dan Du ₄ , Yuehe Lin ₄ , Yong Tang _{1,

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Affiliation

Using an enzyme-linked immunosorbent assay (ELISA) and limited dilution methods to screen and clone antigen-specific hybridoma cells is extremely time-consuming and labor-intensive. This work features a simple and rapid cell surface fluorescence immunosorbent assay (CSFIA), designed for the detection and isolation of antigen-specific hybridoma clones. In this assay, antigens are first anchored to the hybridoma cell surface through a dual-functioning molecular Oleyl-PEG4000-NHS. Specific antibodies secreted from hybridoma cells are then captured by the antigens on the cell surface. Positive hybridoma cells are stained using a fluorescently labeled anti-mouse IgG-Fc antibody. After the addition of a methylcellulose semisolid medium, positive clones are easily picked using a pipet. These positive cell clones can be used to produce monoclonal antibodies after direct expansion. Using this method, positive hybridoma clones against both malachite green and porcine epidemic diarrhea virus are selected with high efficiency. Compared to the ELISA-based method, the CSFIA-based method achieved the capability of isolating >2-fold more hybridoma clones in <25% of the corresponding processing time. In brief, the CSFIA-based method is highly efficient and inexpensive with a simple and direct operation, which is an excellent candidate method for antigen-specific positive clone isolation in a monoclonal antibody preparation.

中文翻译：

抗原特异性杂交瘤细胞克隆的快速分离方法

使用酶联免疫吸附测定（ELISA）和有限的稀释方法来筛选和克隆抗原特异性杂交瘤细胞非常耗时且费力。这项工作的特点是简单，快速的细胞表面荧光免疫吸附测定（CSFIA），旨在检测和分离抗原特异性杂交瘤细胞克隆。在该测定中，首先通过双功能分子Oley1-PEG4000-NHS将抗原锚定到杂交瘤细胞表面。然后，杂交瘤细胞分泌的特异性抗体被细胞表面的抗原捕获。使用荧光标记的抗小鼠IgG-Fc抗体对阳性杂交瘤细胞进行染色。加入甲基纤维素半固体培养基后，可使用移液器轻松挑选阳性克隆。这些阳性细胞克隆可用于直接扩增后产生单克隆抗体。使用这种方法，可以高效地选择针对孔雀石绿和猪流行性腹泻病毒的阳性杂交瘤克隆。与基于ELISA的方法相比，基于CSFIA的方法在不到25％的相应处理时间内即可分离出2倍以上的杂交瘤细胞。简而言之，基于CSFIA的方法高效，便宜，操作简单直接，是单克隆抗体制剂中抗原特异性阳性克隆分离的极佳候选方法。基于CSFIA的方法可在不到25％的相应处理时间内分离出2倍以上的杂交瘤细胞克隆。简而言之，基于CSFIA的方法高效，便宜，操作简单直接，是单克隆抗体制剂中抗原特异性阳性克隆分离的极佳候选方法。基于CSFIA的方法可在不到25％的相应处理时间内分离出2倍以上的杂交瘤细胞克隆。简而言之，基于CSFIA的方法高效，便宜，操作简单直接，是单克隆抗体制剂中抗原特异性阳性克隆分离的极佳候选方法。

更新日期：2018-01-17

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