Label-Free Detection of Mitochondrial Distribution in Cells by Nonresonant Raman Microspectroscopy

Abstract

High spatial resolution Raman maps of fixed cells in an aqueous environment are reported. These maps were obtained by collecting individual Raman spectra via a Raman microspectrometer in a raster pattern on a 0.5-microm grid and assembling pseudocolor maps from the spectral hypercubes by multivariate methods. The Raman maps show the nucleus and the nucleoli of cells as well as subcellular organization in the cytoplasm. In particular, the distribution of mitochondria in the perinuclear region could be demonstrated by correlating distinct areas of the Raman maps with corresponding areas of fluorescence maps of the same cells after staining with mitochondria-specific labels. To the best of our knowledge, this is the first report of label-free detection of mitochondria inside a somatic mammalian cell using Raman microspectroscopy.

Molecular pathology via IR and Raman spectral imaging

“…The last section presented here introduces another aspect of spectroscopic imaging technology that was developed during the past few years [34,35]. This method uses Raman micro-spectroscopy of cells, in a way similar to that described in Section 4.5, but at a much higher spatial resolution.…”

“…In other studies, this methodology was used to distinguish the nucleus, the nucleoli, mitochondriarich and membrane-rich structures inside cells without the use of any contrast agents or dyes [34]. Yet, the most important application of this technology is to monitor transport processes into cells [100,101], for example, the uptake of drug-loaded nanoparticles.…”

“…This results in images which can differentiate the individual cellular components such as the membrane, the cytoplasm, the nucleus and nucleoli, membrane-rich structures in the cytoplasm, mitochondria, etc. [34][35][36]. This method can be viewed as a label-free analog of confocal fluorescence microspectroscopy, and can produce images with similarly high spatial resolution.…”

Spectroscopic and chemometric approaches to radiobiological analyses

“…Quantitative spectral changes attributable to the biochemical processes occurring during cell culture and mitosis [32,33], proliferation [34], differentiation, [35,36], adhesion [37], death [38][39][40][41][42], and invasion [43] have previously been observed with FTIRM and CRM. The increased resolution in CRM imaging has also been exploited to enable the detection of cellular mitochondrial distribution [44] and phagosomes [45]. Early applications of both techniques in radiobiology had focussed on studies with Raman spectroscopy of radiation-induced peroxidation in model phospholipid and membrane systems [46,47], structural changes in protein [48] and in nucleic acids [49].…”

High throughput toxicity screening and intracellular detection of nanomaterials

“…Label‐free imaging of cell organelles,42, 43 uptake and intracellular fate of drug carriers44, 45, 46 and NMs inside individual cells32, 47, 48 have been studied.…”