The Fingerprint of Anti-Bromodeoxyuridine Antibodies and Its Use for the Assessment of Their Affinity to 5-Bromo-2'-Deoxyuridine in Cellular DNA under Various Conditions.
The Fingerprint of Anti-Bromodeoxyuridine Antibodies and Its Use for the Assessment of Their Affinity to 5-Bromo-2'-Deoxyuridine in Cellular DNA under Various Conditions.
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DOI：
10.1371/journal.pone.0132393
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发表时间：
2015
期刊：
影响因子：
3.7
通讯作者：
Koberna K
中科院分区：
文献类型：
--
作者：
Ligasová A;Liboska R;Rosenberg I;Koberna K
我们开发了一种用于分析抗溴脱氧尿核苷抗体亲和力的简单系统。该体系是基于三个不同位置含有5-溴-2‘-脱氧尿苷(BrdU)的锚定寡核苷酸。它允许在固定和通透性细胞中可靠地估计抗溴脱氧尿苷单抗的特定克隆与BrdU的反应性。使用寡核苷酸探针和四种不同的方法检测细胞DNA中的BrdU，我们鉴定了两个抗体克隆，表明在所有测试的方法中对BrdU有足够的反应性。其中一个克隆对5-碘-2‘-脱氧尿苷(IDU)的反应活性高于对BrdU的反应活性。这使我们能够提高所用方案的敏感性，而不会对细胞生理产生负面影响，因为IDU的细胞毒性与BrdU相当，与5-乙炔基-2‘-脱氧尿苷相比可以忽略不计。IDU和改进的DNA氧化降解方法的结合为DNA低降解和高BrdU信号的情况提供了一种敏感和可靠的方法。
We have developed a simple system for the analysis of the affinity of anti-bromodeoxyuridine antibodies. The system is based on the anchored oligonucleotides containing 5-bromo-2'-deoxyuridine (BrdU) at three different positions. It allows a reliable estimation of the reactivity of particular clones of monoclonal anti-bromodeoxyuridine antibodies with BrdU in fixed and permeabilized cells. Using oligonucleotide probes and four different protocols for the detection of BrdU incorporated in cellular DNA, we identified two antibody clones that evinced sufficient reactivity to BrdU in all the tested protocols. One of these clones exhibited higher reactivity to 5-iodo-2'-deoxyuridine (IdU) than to BrdU. It allowed us to increase the sensitivity of the used protocols without a negative effect on the cell physiology as the cytotoxicity of IdU was comparable with BrdU and negligible when compared to 5-ethynyl-2'-deoxyuridine. The combination of IdU and the improved protocol for oxidative degradation of DNA provided a sensitive and reliable approach for the situations when the low degradation of DNA and high BrdU signal is a priority.
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影响因子：
3.7
作者：
Ligasová A;Strunin D;Liboska R;Rosenberg I;Koberna K
通讯作者：
Koberna K
影响因子：
4.7
作者：
Hong, Vu;Steinmetz, Nicole F.;Manchester, Marianne;Finn, M. G.
通讯作者：
Finn, M. G.
影响因子：
3.7
作者：
Ligasova, Anna;Strunin, Dmytro;Koberna, Karel
通讯作者：
Koberna, Karel
影响因子：
5.3
作者：
Barbie, DA;Kudlow, BA;Kennedy, BK
通讯作者：
Kennedy, BK
影响因子：
3.9
作者：
Ross, Heather H.;Rahman, Maryam;Levkoff, Lindsay H.;Millette, Sebastien;Martin-Carreras, Teresa;Dunbar, Erin M.;Reynolds, Brent A.;Laywell, Eric D.
通讯作者：
Laywell, Eric D.