Long-Term Real-Time Tracking of Lanthanide Ion Doped Upconverting Nanoparticles in Living Cells
Long-Term Real-Time Tracking of Lanthanide Ion Doped Upconverting Nanoparticles in Living Cells
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DOI：
10.1002/anie.201007979
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发表时间：
2011-01-01
影响因子：
16.6
通讯作者：
Suh, Yung Doug
中科院分区：
文献类型：
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作者：
Nam, Sang Hwan;Bae, Yun Mi;Suh, Yung Doug
近年来，将纳米粒子用于各种生物应用引起了人们的极大兴趣。[1]纳米粒子可以通过可控的方式合成，使其具有理想的尺寸、形状和光学或磁性。此外，人们还可以通过化学表面修饰和配体的偶联来提供具有生物功能的纳米颗粒。[2]纳米颗粒的这种内在和外在性质使其能够作为优秀的生物成像探针和细胞水平的诊断/治疗试剂。[3]在迄今为止发展起来的各种纳米颗粒体系中，半导体纳米晶或量子点(Qds)的应用最为广泛。量子点是非常明亮和光稳定的，并且表现出非常好的光谱特性(即宽吸收和窄发射带)，适合于多色检测。[4-6]然而，诸如光闪烁、[7]无辐射暗粒子的存在、[8]和潜在的细胞毒性[9]等缺点限制了它们的应用。近年来，几种不同类型的发光纳米颗粒已被引入生物应用。例如，具有氮空位中心的纳米钻石(NDS)被发现具有高度的光致发光，同时不表现出光闪烁和光漂白，[10，11]，甚至可用作超分辨率光学显微镜的成像探针。[12]然而，将NDS应用于生物成像具有局限性，特别是在长期跟踪研究的情况下，因为蓝色或绿色区域(通常为488或532 nm)的激发可能导致对细胞的致命光损伤或对组织的低渗透深度。相比之下，单壁碳纳米管(SWNTs)被证明适合于生物成像，因为它的激发和发射位于近红外(NIR)光谱范围内。[13-15]然而，由于SWNTs通常长于100 nm，因此被认为太大而不能用作生物标记。与此同时，稀土离子掺杂的上转换纳米粒子(UCNPs)由于其独特的光学性质而引起了人们的极大关注。首先，近红外激发的双光子上转换为可见光子的发射是如此有效，以至于输出数十毫瓦的微小连续波(CW)半导体激光器(980 Nm)足以作为激发源。[16，17]其次，通过使用近红外激发，人们可以抑制细胞的自发荧光，对活细胞的光损伤很小，并实现相对深入的组织渗透。最后，UCNP在毫秒和秒时间尺度上既不表现出光闪烁，也不表现出光漂白，即使连续激发几个小时，[20，21]它们的细胞毒性非常低，[20，21]。[20，23]因此，UCNPs成为最有前途的生物成像纳米粒子系统之一，并通过设计新的合成策略来改进其性能(例如，增加发光强度和减小颗粒尺寸)。[24]在此，我们首次报道了在活细胞中对UCNPs进行的实时跟踪研究。由于UCNPs显著的光稳定性和近红外激发的非侵入性，我们能够不间断地观察UCNPs在细胞内长达6h的运动。我们首先评估了使用NIR辐射作为激发源的好处，以论证使用UCNPs进行长期活细胞成像的可行性。Yb3+和…共掺六方相NaYF4
Recently, there has been great interest in employing nanoparticles for various biological applications.[1] Nanoparticles can be synthesized in a controlled manner such that they have desirable sizes, shapes, and optical or magnetic properties. In addition, one may provide nanoparticles with biological functions through chemical surface modifications and conjugation of ligands.[2] Such intrinsic and extrinsic properties of nanoparticles enable them to be used as excellent biological imaging probes and diagnostic/therapeutic agents at the cellular level.[3] Among the various nanoparticle systems developed thus far, semiconductor nanocrystals or quantum dots (QDs) are most widely used. QDs are extremely bright and photostable, and exhibit excellent spectral properties (ie, broad absorption and narrow emission bands) suited for multicolor detection.[4–6] However, the drawbacks such as photoblinking,[7] the presence of nonradiant dark particles,[8] and potential cytotoxicity [9] limit their applicability. In recent years, several alternative types of luminescent nanoparticles have been introduced for biological applications. For example, nanodiamonds (NDs) with nitrogen vacancy centers were found to be highly photoluminescent while exhibiting no photoblinking and photobleaching,[10, 11] and even useful as the imaging probe for super-resolution optical microscopy.[12] However, applying NDs for biological imaging has limitations, especially in the case of long-term tracking studies, since the excitation in the blue or green region (typically 488 or 532 nm) might result in fatal photodamage to cells or low penetration depth into tissues. In contrast, single-walled carbon nanotubes (SWNTs) were shown to be appropriate for biological imaging in that the excitation and emission lie in the near-infrared (NIR) spectral range.[13–15] However, being longer than 100 nm typically, SWNTs are considered to be too large to be used as biolabels. Meanwhile, lanthanide ion doped upconverting nanoparticles (UCNPs), which emit in the visible range upon absorption of NIR photons, have attracted great attention owing to their unique optical properties. First, two-photon upconversion of NIR excitation to the emission of a visible photon is so efficient that a tiny continuous-wave (CW) diode laser (980 nm) with the output of tens of milliwatts is sufficient as the excitation source.[16, 17] Second, by employing NIR excitation, one can suppress cellular autofluorescence, induce little photodamage to living cells, and achieve relatively deep penetration into tissues.[18, 19] Finally, UCNPs exhibit neither photoblinking on the millisecond and second time scales nor photobleaching even with hours of continuous excitation,[20, 21] their cytotoxicity is very low,[20, 22] and the inclusion or doping of Gd3+ ions in the host materials endows UCNPs with an additional modality for magnetic resonance imaging (MRI).[20, 23] As a result, UCNPs became one of the most promising nanoparticle systems for biological imaging and there are continuing efforts to improve their properties (eg, increasing luminescence intensity and reducing the particle size) by designing new synthetic strategies.[24] Herein, we report the first real-time tracking study with UCNPs at the single vesicle level in living cells. Thanks to the remarkable photostability of UCNPs and the noninvasiveness of NIR excitation, we were able to visualize the intracellular movements of UCNPs for as long as 6 h without interruption.We first assessed the benefits of using NIR radiation as the excitation source to demonstrate the feasibility of long-term live-cell imaging with UCNPs. The UCNPs (hexagonal-phase NaYF4 co-doped with Yb3+ and …