IDENTIFICATION OF THE CHICK NEURAL RETINA CELL-SURFACE N-ACETYLGALACTOSAMINYLTRANSFERASE USING MONOCLONAL-ANTIBODIES
IDENTIFICATION OF THE CHICK NEURAL RETINA CELL-SURFACE N-ACETYLGALACTOSAMINYLTRANSFERASE USING MONOCLONAL-ANTIBODIES
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DOI：
10.1002/jcb.240320205
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发表时间：
1986-01-01
影响因子：
4
通讯作者：
LILIEN, J
中科院分区：
文献类型：
--
作者：
BALSAMO, J;PRATT, RS;LILIEN, J
完整的鸡胚胎视网膜神经细胞表面有一个N-乙酰氨基半乳糖转移酶，该酶催化来自UDP-N-乙酰氨基半乳糖的N-乙酰氨基半乳糖掺入内源性大分子受体。在用Triton X-100处理膜富集组分后，该酶及其内源受体可以被分离为颗粒复合体。在本文中，我们报道了两种产生单抗的融合：一种是以颗粒复合体作为免疫原，另一种是以组织培养条件培养液中发现的缺乏内源性受体活性的可溶性N-乙酰氨基半乳糖转移酶作为免疫原。来自两种融合的抗体识别与颗粒转移酶/受体复合体密切相关的抗原和具有N-乙酰半乳糖基转移酶活性的可溶性抗原。该抗体识别CA-MR 220,000的一个组分，经SDS-凝胶电泳法显示N-乙酰氨基半乳糖转移酶活性，并将其转移到硝基纤维素。该组分在双向凝胶电泳法上与一种可碘化的细胞表面组分共存，该组分在细胞表面的存在与内源性转移酶的活性有关。我们得出的结论是，抗体识别转移酶本身。免疫组织化学分析表明，该酶最初定位于胚胎神经视网膜的整个细胞表面，但在成年后仅限于外丛状层和外节。
Intact embryonic chick neural retina cells have at their surface an N-acetylgalactosaminyltransferase which catalyzes the incorporation of N-acetylgalactosamine from UDP-N-acetylgalactosamine into endogenous macromolecular acceptors. The enzyme along with its endogenous acceptors can be isolated as a particulate complex following treatment of membrane-enriched fractions with Triton X-100. In this paper we report on two separate fusions generating monoclonal antibodies: one using as immunogen the particulate complex and the second using as immunogen a soluble N-acetylgalactosaminyltransferase found in tissue-culture-conditioned medium which lacks endogenous acceptor activity. Antibodies from both fusions recognize an antigen which is tightly associated with the particulate transferase/acceptor complex and a soluble antigen having N-acetylgalactosaminyltransferase activity toward exogenously added acceptors. The antibodies recognize a component of ca Mr 220,000, which shows N-acetylgalactosaminyltransferase activity after SDS-gel electrophoresis and transfer to nitrocellulose. This component comigrates on two-dimensional gel electrophoresis with an iodinatable cell surface component whose presence at the cell surface correlates with endogenous transferase activity. We conclude that the antibodies recognize the transferase enzyme itself. Immunohistochemical analysis shows that the enzyme is initially localized throughout the embryonic neural retina in a pattern indicative of a cell surface disposition but becomes restricted to the outer plexiform layer and to outer segments in the adult.