靶向卵巢癌细胞荧光-磁共振双模态纳米探针的制备及细胞成像

摘要

目的 构建靶向卵巢癌细胞的荧光-磁共振双模态纳米探针锰掺杂-碳量子点(Mn-CDs)@抗-人附睾蛋白4(HE4)单克隆抗体(Mn-CDs@Anti-HE4 mAb),探讨其用于荧光成像及MRI 的潜能.方法 先采用溶剂热法在150℃下制备Mn-CDs,检测其平均粒径、荧光成像性能及MRI性能.用新型四唑化合物(MTS)法检测Mn-CDs对人卵巢癌干细胞HO-8910及人脐静脉血管内皮细胞EA.hy926的细胞毒性.以缩合反应构建Mn-CDs@Anti-HE4 mAb,用紫外光谱进行表征,并进行细胞水平的荧光成像及MRI,评估其靶向性.采用单因素方差分析和最小显著差异t检验处理数据.结果 制得球形Mn-CDs,粒径为(4.64±0.85)nm,最佳激发波长为360 nm,最大发射波长为440 nm,T1弛豫效率为(3.26±0,04)mmol.L-1·s-1.HO-8910细胞、EA.hy926细胞在不同质量浓度Mn-CDs 作用下存活率差异均有统计学意义(F=t 947.509、260.174,均P<0.05);但Mn-CDs质量浓度在0～2.5 mg/ml时,对HO-8910细胞及EA.hy926细胞均无毒性(均P>0.05);而在3.0-4.5 mg/ml时,随着 Mn-CDs质量浓度递增,2种细胞存活率均递减(P<0.05).Mn-CDs@Anti-HE4 mAb在荧光成像及 MRI中均能靶向识别HO-8910细胞.结论成功构建的Mn-CDS@Anti-HE4 mAb具有用于荧光成像、MRI的潜能及靶向识别能力,有可能为卵巢癌的早期诊断提供新手段.%Objective To fabricate manganese-doped carbon quantum dots(Mn-CDs)@anti-human epididymis protein 4(HE4)monoclonal antibody(Mn-CDs@Anti-HE4 mAb)dual-modal fluorescent-magnetic nanoprobe for ovarian cancer cells targeting imaging,and evaluate its potential on fluorescent imaging and MRL Methods Mn-CDs were synthesized at 150 ℃ with solvothermal method.The average diameter,fluorescent capability and MRI efficiency were determined.The cytotoxicity of Mn-CDs in vitro was evaluated by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium(MTS)assay with HO-8910 ovarian cancer stem cells and EA.hy926 human umbilical vein endothelial cells.Mn-CDs@Anti-HE4 mAb was fabricated with condensation reaction and characterized by ultraviolet(UV)absorption spectra.Fluorescence imaging and MRI in vitro was performed for cancer cell-targeting study.One-way analysis of variance and the least significant difference t test were used to analyze the data.Results The Mn-CDs with diameter of(4.64±0.85)nm showed a well-defined spherical morphology.The fluorescent spectra of Mn-CDs exhibited a typical excitation-dependent behavior with an excitation maximum at 360 nm and emission maximum at 440 nm.The T1 relaxation rate was(3.26±0.04)mmol ? L-1 ? s-1.The cytotoxicity tests in vitro showed that the survival rates of HO-8910 cells and EA.hy926 cells were both significantly different after treated with different concentrations of Mn-CDs(F= 1 947.509,260.174,both P<0.05),and there was no cytotoxicity in both HO-8910 cells and EA.hy926 cells at concentrations of MnCDs within 0-2.5 mg/ml(all P>0.05),while the survival rates of the two kinds of cells were descended with the increasing of concentration within 3.0-4.5 mg/ml(P<0.05).Mn-CDs@Anti-HE4 mAb could target HO-8910 cells on fluorescence imaging and MRI.Conclusions Mn-CDs@Anti-HE4 mAb,with good potential on fluorescence imaging,MRI and targeting ability,is successfully synthesized.It may provide a new method for early diagnosis of ovarian cancer.