Ling Crystal Chia Yin, Fullwood Melissa Jane
School of Biological Sciences, Nanyang Technological University, Singapore, Singapore.
PLoS One. 2024 Dec 27;19(12):e0316359. doi: 10.1371/journal.pone.0316359. eCollection 2024.
Immunofluorescence is highly dependent on antibody-antigen interactions for accurate visualization of proteins and other biomolecules within cells. However, obtaining antibodies with high specificity and affinity for their target proteins can be challenging, especially for targets that are complex or naturally present at low levels. Therefore, we developed AptaFluorescence, a protocol that utilizes fluorescently labeled aptamers for in vitro biomolecule visualization. Aptamers are single-stranded nucleic acid molecules that fold into three-dimensional structures to bind biomolecules with high specificity. AptaFluorescence targeting the c-MYC protein was evaluated in doxycycline-inducible c-MYC U2OS cells. AptaFluorescence signals were more distinct compared to the diffuse immunofluorescence signals. AptaFluorescence also reliably differentiated doxycycline-treated cells from untreated cells. In conclusion, AptaFluorescence is a novel, easy to perform, aptamer-based protocol that will have broad applicability across various biological endeavours for visualizing biomolecules, especially in cases where antibodies with high affinity and specificity for their target proteins are lacking.
免疫荧光高度依赖抗体 - 抗原相互作用来准确可视化细胞内的蛋白质和其他生物分子。然而，获得对其靶蛋白具有高特异性和亲和力的抗体可能具有挑战性，特别是对于复杂或天然存在水平较低的靶标。因此，我们开发了AptaFluorescence，这是一种利用荧光标记的适体进行体外生物分子可视化的方案。适体是单链核酸分子，可折叠成三维结构以高特异性结合生物分子。在强力霉素诱导的c-MYC U2OS细胞中评估了靶向c-MYC蛋白的AptaFluorescence。与弥漫性免疫荧光信号相比，AptaFluorescence信号更清晰。AptaFluorescence还能可靠地区分经强力霉素处理的细胞和未处理的细胞。总之，AptaFluorescence是一种新颖、易于操作的基于适体的方案，将在各种生物研究中广泛应用于生物分子可视化，特别是在缺乏对其靶蛋白具有高亲和力和特异性的抗体的情况下。
