Date Published: December 31, 2017
Publisher: John Wiley and Sons Inc.
Author(s): Meysam Keshavarz, Bo Tan, Krishnan Venkatakrishnan.
Herein, a label‐free multiplex photoluminescent silicon nanoprobe (PLSN‐probe) is introduced as a potential substitute for quantum dots (QDs) in bioimaging. An inherently non‐photoluminescent silicon substrate is altered to create the PLSN‐probe, to overcome the major drawbacks of presently available QDs. Additionally, crystallinity alterations of the multiplane crystalline PLSN‐probes lead to broad absorption and multiplex fluorescence emissions, which are attributed to the simultaneous existence of multiple crystal planes. The PLSN‐probe not only demonstrates unique optical properties that can be exploited for bioimaging but also exhibits cell‐selective uptake that allows the differentiation and diagnosis of HeLa and fibroblast cells. Moreover, multiplex emissions of the PLSN‐probe illuminate different organelles such as the nucleus, nucleolemma, and cytoskeleton, depending on size‐based preferential uptake by the cell organs. This in vitro study reveals that cancerous HeLa cells have a higher propensity for taking up the PLSN‐probe compared to fibroblast cells, allowing the diagnosis of cancerous HeLa cells. Additionally, the fluorescence intensity per unit area of the cell is found to be a reliable means for distinguishing between dead and healthy cells. It is anticipated that the multifunctionality of the PLSN‐probes will lead to better insight into the use of such probes for bioimaging and diagnosis applications.
Fluorescence (FL) microscopy is extensively used as a crucial tool for biological applications such as biomedical imaging and clinical diagnostics.1 However, conventional cell imaging techniques that utilize organic fluorophores have some major problems due to their nonspecific accumulation within cells and photobleaching of the dyes.2, 3 Commercially available organic dyes have been routinely used in life sciences for FL imaging; however, these dyes have some inadequacies that hinder them in the rapidly growing field of bioimaging. The main drawbacks include a limited lifetime (a few nanoseconds), progressive reduction of FL (photobleaching), low contrast in some applications, emission spectra with a red tail, and low emission times. Hence, an imaging method that is not reliant on fluorophore dyes is needed.4 Therefore, extensive efforts have been made to produce an alternative to organic fluorescent molecules to alleviate these problems.5
Here, we presented a novel approach using polyhedral PLSN‐probes with a broad range of absorption and long Stokes shifts that exhibited multiplex emissions. This label‐free method has shown a precise capability of cell uptake that strongly improves upon the lengthy labeling process of other methods. However, this attribute was found to be cell‐selective, i.e., the HeLa cells demonstrated significantly extended internalization, whereas the fibroblast cells showed a weaker tendency to uptake the PLSN‐probes. The multicrystallinity of the polyhedral structure allows higher absorption for the PLSN‐probe, and hence, a greater FL emission was observed. The PLSN‐probes demonstrated excellent dispersion in aqueous cell culture medium and systematic division into smaller parts that internalize into different organs of the cell, including the nucleus, nucleolemma, and cytoskeleton by which 3D visualization of intercellular organs of the cell is made possible. Moreover, we found that the FL intensity of the PLSN‐probes per cell area increases upon shrinkage of the cytoskeleton of the cell. Hence, this characteristic allows one to conclusively differentiate healthy cells from dead cells. In addition, the proposed probes can be used to diagnose HeLa cells as well as to screen dead cells. Further investigations are required to study the interaction of different cell lines with these PLSN‐probes.
To investigate the morphology and dimensions of the synthesized PLSN‐probes, a JEOL JSM‐4800 field emission scanning electron microscope and a JEOL JEM 2100 HR‐TEM were used. HR‐TEM was performed to determine the size distribution and morphology of the PLSN‐probes; for this method, a droplet of suspended PLSN‐probes was placed on a copper grid for direct observation after drying. Image analysis software was used to analyze the HR‐TEM images, perform size measurements, and compile a distribution histogram. An HR‐TEM diffraction pattern (FFT) was also obtained to confirm the formation of crystalline PLSN‐probes. Energy‐dispersive X‐ray spectroscopy (EDX) analyses using Oxford Instruments further corroborated the formation of pure PLSN‐probes and the elemental mapping evidenced the self‐internalization of the synthesized PLSN‐probes into the cells. The Raman spectra were recorded with a Bruker SENTERRA dispersive Raman microscope using a wavelength of 325 nm. The Raman spectra showed a strong band and sharp peaks at 521 cm−1, which are characteristics of crystalline PLSN‐probes. UV–vis–NIR absorption and reflection spectra were obtained with a Hitachi UV‐3100 UV–vis–NIR spectrophotometer. PL spectra were recorded on a Shimadzu RF‐5301PC spectrofluorophotometer at room temperature.
The authors declare no conflict of interest.