Evaluation of fluorescence-based viability stains in cells dissociated from scleractinian coral Pocillopora damicornis.
Evaluation of fluorescence-based viability stains in cells dissociated from scleractinian coral Pocillopora damicornis.
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DOI：
10.1038/s41598-022-19586-7
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发表时间：
2022-09-12
影响因子：
4.6
通讯作者：
Lewinski, Nastassja A.
中科院分区：
文献类型：
--
作者：
Roger, Liza M.;Darko, Yaa Adarkwa;Bernas, Tytus;White, Frances;Olaosebikan, Monsurat;Cowen, Lenore;Klein-Seetharaman, Judith;Lewinski, Nastassja A.
建立细胞活力测定，如常用的台盼蓝染色法珊瑚细胞的应用程序是不简单的，由于不同的文化参数和不同的细胞功能，具体到哺乳动物细胞相比，海洋无脊椎动物。使用Pocillopora damicornis作为模型，我们的特点是自发荧光和测试不同的荧光染料对组合，以确定替代的生存力指标。在暴露24小时后，使用荧光染料对Hoechst 33342和SYTOX橙子测量不同代表性分子（即小有机分子、蛋白质和纳米颗粒（NP））的细胞毒性。我们的研究结果表明，这种染料对可以清楚地测量荧光蛋白和叶绿素的存在下。将红角裸腹蛛细胞分别暴露于浓度范围为0.5至100 µg/mL的Triton-X100、胰岛素或二氧化钛（TiO 2）纳米颗粒24 h，显示Triton-X100的LC 50为0.46 µg/mL，TiO 2纳米颗粒为6.21 µg/mL，胰岛素为33.9 µg/mL。这项工作提出了用于定制染料对膜完整性为基础的细胞活力测定考虑的物种和基因型特异性自发荧光的石珊瑚，即：内源性荧光表征，然后选择不与内源性信号重叠的染料的方法。
The application of established cell viability assays such as the commonly used trypan blue staining method to coral cells is not straightforward due to different culture parameters and different cellular features specific to mammalian cells compared to marine invertebrates. Using Pocillopora damicornis as a model, we characterized the autofluorescence and tested different fluorescent dye pair combinations to identify alternative viability indicators. The cytotoxicity of different representative molecules, namely small organic molecules, proteins and nanoparticles (NP), was measured after 24 h of exposure using the fluorescent dye pair Hoechst 33342 and SYTOX orange. Our results show that this dye pair can be distinctly measured in the presence of fluorescent proteins plus chlorophyll. P. damicornis cells exposed for 24 h to Triton-X100, insulin or titanium dioxide (TiO2) NPs, respectively, at concentrations ranging from 0.5 to 100 µg/mL, revealed a LC50 of 0.46 µg/mL for Triton-X100, 6.21 µg/mL for TiO2 NPs and 33.9 µg/mL for insulin. This work presents the approach used to customize dye pairs for membrane integrity-based cell viability assays considering the species- and genotype-specific autofluorescence of scleractinian corals, namely: endogenous fluorescence characterization followed by the selection of dyes that do not overlap with endogenous signals.
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影响因子：
4.6
作者：
Klenow MB;Heitmann ASB;Nylandsted J;Simonsen AC
通讯作者：
Simonsen AC
影响因子：
6.8
作者：
Dayeh, VR;Chow, SL;Bols, NC
通讯作者：
Bols, NC
影响因子：
3.7
作者：
Alieva NO;Konzen KA;Field SF;Meleshkevitch EA;Hunt ME;Beltran-Ramirez V;Miller DJ;Wiedenmann J;Salih A;Matz MV
通讯作者：
Matz MV
DOI：
10.1007/s10126-021-10031-w
发表时间：
2021-06
期刊：
Marine biotechnology (New York, N.Y.)
影响因子：
--
作者：
Kawamura K;Nishitsuji K;Shoguchi E;Fujiwara S;Satoh N
通讯作者：
Satoh N
影响因子：
2.9
作者：
Bessette, PH;Daugherty, PS
通讯作者：
Daugherty, PS