Development of signal multiplication system for quinone linked immunosorbent assay (Multi-QuLISA) by using poly-L-lysine dendrigraft and 1,2-naphthoquinone-4-sulfonate as enzyme-free tag
Development of signal multiplication system for quinone linked immunosorbent assay (Multi-QuLISA) by using poly-L-lysine dendrigraft and 1,2-naphthoquinone-4-sulfonate as enzyme-free tag
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DOI：
10.1016/j.talanta.2022.123911
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发表时间：
2022-09-19
期刊：
影响因子：
6.1
通讯作者：
Kuroda, Naotaka
中科院分区：
文献类型：
--
作者：
Kaladari, Fatema;El-Maghrabey, Mahmoud;Kuroda, Naotaka
开发了一种灵敏且稳定的信号倍增醌联免疫吸附测定法 (Multi-QuLISA)。在 Multi-QuLISA 中，使用与生物素连接的寡聚醌，即生物素-8mer-萘醌 (Bio8mer-NQ) 作为信号生成标记。 Bio8mer-NQ 由第 1 代聚-L-赖氨酸树状移植物 (DPLL G1) 形成，这是一种受控支化寡聚物，由八个赖氨酸部分和九个游离氨基作为主链组成。 DPLL G1 的九个氨基之一连接到生物素部分，而其他八个连接到 1,2-萘醌-4-磺酸盐 (NQS)。 Bio8mer-NQ 使用抗生物素蛋白作为辅助结合剂标记生物素化检测抗体。然后，Bio8mer-NQ 中的多醌与二硫苏糖醇和鲁米诺经历氧化还原循环，产生强烈的化学发光。标准 ELISA 使用的标记酶在不同条件下易受影响且稳定性较差。 Bio8mer-NQ在不同的剧烈pH和温度条件、水解酶等条件下表现出比酶（生物素-HRP）更好的稳定性。此外，Bio8mer-NQ基于醌的氧化还原循环同时用作化学发光和比色标记，其LOD分别为1.5和6.5 nM。该方法可以在内部设计的免疫测定中检测低至 0.2 nM 的生物素化免疫复合物，其灵敏度比生物素 HRP 大约高 25 倍。最终，Bio8mer-NQ 成功应用于 Multi-QuLISA 检测 β-酪蛋白，灵敏度为 3.2 ng/mL，而传统 ELISA 的 LOD 为 35 ng/mL。总体而言，Bio8mer-NQ 是一种稳定的化合物，可用作免疫测定中酶的出色替代品，并且可用于比色和化学发光测定，具有良好的灵敏度。
A sensitive and stable signal multiplied quinone-linked immunosorbent assay (Multi-QuLISA) was developed. In Multi-QuLISA, an oligomerized quinone linked to biotin, namely biotin-8mer-naphthoquinone (Bio8mer-NQ), is used as a signal-generating label. Bio8mer-NQ is formed from a dendrigraft poly -L-lysine generation 1 (DPLL G1), a controlled branched oligomer composed of eight lysine moieties with nine free amino groups as a backbone. One of the nine amino groups of DPLL G1 is attached to biotin moiety, while the other eight are attached to 1,2-naphthoquinone-4-sulfonate (NQS). Bio8mer-NQ labels a biotinylated detection antibody using avidin as a co -binder. Then, multi-quinones in Bio8mer-NQ undergo a redox cycle with dithiothreitol and luminol, gener-ating strong chemiluminescence. Standard ELISA uses a label enzyme that suffers from vulnerability in different conditions and poor stability. Bio8mer-NQ showed better stability than the enzyme (biotin-HRP) under different drastic pH and temperature conditions, hydrolytic enzymes, etc. Furthermore, Bio8mer-NQ was used as both chemiluminescence and colorimetric label based on the redox cycle of quinone, and it had LODs of 1.5 and 6.5 nM, respectively. The method could detect biotinylated immunocomplex in an in-house designed immunoassay down to 0.2 nM, which is about 25 times more sensitive than biotin HRP. Eventually, Bio8mer-NQ was applied successfully in Multi-QuLISA for detecting beta-casein with a sensitivity of 3.2 ng/mL, while the conventional ELISA had an LOD of 35 ng/mL. Overall, Bio8mer-NQ is a stable compound that could be used as an excellent replacement for the enzyme in immunoassay and can be used in both colorimetric and chemiluminescence assays with good sensitivity.