Improved high-affinity monoclonal antibody to iododeoxyuridine.
Improved high-affinity monoclonal antibody to iododeoxyuridine.
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改良的碘脱氧尿苷高亲和力单克隆抗体。
DOI：
10.1002/cyto.990070602
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发表时间：
1986
期刊：
影响因子：
--
通讯作者：
Stanker,L
中科院分区：
文献类型：
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作者：
Vanderlaan,M;Watkins,B;Thomas,C;Dolbeare,F;Stanker,L
用酶免疫分析法(EL ISA)和流式细胞术对3种单抗(Iu-1、Iu-4和B-44)识别溴脱氧尿苷(BrdUrd)和碘脱氧尿苷(IdUrd)取代的DNA、核苷酸和核苷的能力进行了评价。Iu-4是一种新的单抗，由5-碘-2‘-脱氧尿苷-5’-单磷酸(IdUMP)通过磷酸连接到白蛋白免疫小鼠而来。该免疫原在DNA中与IdUMP相似。在竞争ELISA检测中，IU-4偏好IdUrd而不是BrdUrd，并且偏好卤代核苷酸而不是相应的核苷。在ELISA和Flow分析中，Iu-4识别DNA中的IdUrd的替换频率至少低至每1000个正常碱基中有一个IdUrd。IdUrd-DNA与IdUrd-DNA的高亲和力与Iu-1和B-44形成鲜明对比，Iu-1和B-44对碱基取代的频率表现出很强的结合依赖性，并且在Flow和ELISA法中都需要基本上完全被BrdUrd取代的DNA才能结合。Iu-4与DNA中IdUrd的高亲和力和IdUrd残基间距的独立性使其成为全细胞内定量标记卤代胸苷类似物的优良试剂。
Three monoclonal antibodies (Mabs), IU‐1, IU‐4, and B‐44 were evaluated in enzyme immunoassays (ELISA) and by flow cytometry for their abilities to recognize bromodeoxyuridine (BrdUrd)‐ and iododeoxyuridine (IdUrd)‐substituted DNA's, nucleotides, and nucleosides. IU‐4 is a new Mab, derived from mice immunized with 5‐iodo‐2′‐deoxyuridine‐5′‐monophosphate (IdUMP) conjugated through the phosphate group to albumin. This immunogen was selected to resemble IdUMP in DNA. In competition ELISA assays, IU‐4 prefers IdUrd to BrdUrd and prefers halogenated nucleotides over the corresponding nucleosides. In both ELISA and flow analysis, IU‐4 recognizes IdUrd in DNA at substitution frequencies at least as low as one IdUrd one per 1,000 normal bases. The high affinity of IU‐4 for IdUrd‐DNA contrasts with IU‐1 and B‐44, which show a strong binding dependency on the frequency of base substitution and require DNA that is essentially fully substituted with BrdUrd for binding in both flow and ELISA assays. The high affinity of IU‐4 for IdUrd in DNA and its independence of IdUrd residue spacing make it a superior reagent for the quantitative labeling of halogenated thymidine analogues in whole cells.