Growth substrate induced functional changes elucidated by FTIR and Raman spectroscopy in in–vitro cultured human keratinocytes

Meade, Aidan D.; Lyng, Fiona M.; Knief, Peter; Byrne, Hugh J.

doi:10.1007/s00216-006-0876-5

Cited by 109 publications

(120 citation statements)

References 58 publications

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“…Cell culture on such substrates has been demonstrated to be toxic to the cell, although use of molecular biochemical coatings (such as gelatin) have been shown to ameliorate such effects [37]. Cells may be studied in their live form with CRM, but may also be preserved with chemical fixation for both CRM and FTIRM (which will also require sample dessication), and this has been demonstrated to adjust both sets of spectra such that experimental controls preserved in the same manner are generally required [63].…”

Section: Sample Preparation Considerationsmentioning

confidence: 99%

“…Cells were cultured on low emissivity silver oxide coated glass slides (MirrIR, Kevley Technologies), on which a 2% gelatin coating was deposited to enable attachment of the cell and improve cell viability [37]. Doses (ten points) ranging from 0 Gy to 5 Gy were delivered to the samples, and they were fixed in 4% neutral-buffered formalin at time points ranging from 6 hours to 96 hours post-irradiation.…”

Section: Sample Preparation and Data Acquisitionmentioning

confidence: 99%

“…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].…”

Section: Introductionmentioning

confidence: 99%

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Spectroscopic and chemometric approaches to radiobiological analyses

Meade

Byrne²,

Lyng

2010

Mutation Research/Reviews in Mutation Research

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Vibrational spectroscopy is an attractive modality for the analysis of biological samples, providing a complete non-invasive acquisition of the biochemical fingerprint of the sample. It has been demonstrated that this data provides the means to assay multiple functional responses of a biological system at a spatial resolution as low as a micron within the sample. As the interaction of ionizing radiation with biological systems involves chemical reactions between the products of radiation induced damage and various structural and functional units within the cell, the vibrational spectroscopic modalities have received attention as potential measurement platforms for the in-situ examination of the chemistry of biological species in radiobiology. This presents challenges in relation to sample preparation and the construction of suitable analytical methodologies. In this work protocols for sample preparation and approaches to multivariate analysis of vibrational spectra in radiobiological analysis are detailed and the utility of the methodology in analyzing the evolution of biochemical responses to radiobiological damage are highlighted.

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Section: Sample Preparation Considerationsmentioning

confidence: 99%

Section: Sample Preparation and Data Acquisitionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Spectroscopic and chemometric approaches to radiobiological analyses

Meade

Byrne²,

Lyng

2010

Mutation Research/Reviews in Mutation Research

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“…The HaCaT cell line seemed to be more affected by the different substrates and an increase from 34% to 97% was observed, a 285% increase in viability in the collagen gels compared to the uncoated quartz substrates. For comparison, the % increase in viability of HaCaT cells on fibronectin, gelatine and lamanin coated quartz substrates as measured by the Alamar Blue assay was observed to be ~45, 66 and 92 % respectively 19 . Clearly the increased viability affected by the 3-D collagen gel is significantly higher.…”

Section: Cell Viability In Collagen Gel Matricesmentioning

confidence: 99%

“…Improved biocompatibility of quartz for biospectroscopic applications can be achieved by the deposition of a thin layer of gelatin although spectral contributions from the substrate itself are still problematic 19 .…”

Section: Introductionmentioning

confidence: 99%

Three dimensional collagen gels as a cell culture matrix for the study of live cells by Raman spectroscopy

Bonnier¹,

Meade²,

Merzha³

et al. 2010

Analyst

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Three dimensional collagen gels are evaluated as matrices for the study of live cells by Raman spectroscopy. The study is conducted on a human lung adenocarcinoma (A549) and a spontaneously immortalized human epithelial keratinocyte (HaCaT) cell line. It is demonstrated, using the Alamar Blue assay, that both cell models exhibit enhanced viability in collagen matrices compared to quartz substrates, commonly used for Raman spectroscopy. Using principal component analysis, it is shown that the Raman spectral analysis of cells in collagen matrices are minimally contaminated by substrate contributions and the cell to cell spectral variations are greatly reduced compared to those measured on quartz substrates. Furthermore, the spectral measurements are seen to have little contribution from the cell culture medium, implying that cultures can be kept viable over prolonged measurement or mapping procedures.

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Raman spectroscopy in biomedicine – non‐invasive in vitro analysis of cells and extracellular matrix components in tissues

Brauchle

Schenke‐Layland

2012

Biotechnology Journal

119

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Raman spectroscopy is an established laser-based technology for the quality assurance of pharmaceutical products. Over the past few years, Raman spectroscopy has become a powerful diagnostic tool in the life sciences. Raman spectra allow assessment of the overall molecular constitution of biological samples, based on specific signals from proteins, nucleic acids, lipids, carbohydrates, and inorganic crystals. Measurements are non-invasive and do not require sample processing, making Raman spectroscopy a reliable and robust method with numerous applications in biomedicine. Moreover, Raman spectroscopy allows the highly sensitive discrimination of bacteria. Rama spectra retain information on continuous metabolic processes and kinetics such as lipid storage and recombinant protein production. Raman spectra are specific for each cell type and provide additional information on cell viability, differentiation status, and tumorigenicity. In tissues, Raman spectroscopy can detect major extracellular matrix components and their secondary structures. Furthermore, the non-invasive characterization of healthy and pathological tissues as well as quality control and process monitoring of in vitro-engineered matrix is possible. This review provides comprehensive insight to the current progress in expanding the applicability of Raman spectroscopy for the characterization of living cells and tissues, and serves as a good reference point for those starting in the field.

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Growth substrate induced functional changes elucidated by FTIR and Raman spectroscopy in in–vitro cultured human keratinocytes

Cited by 109 publications

References 58 publications

Spectroscopic and chemometric approaches to radiobiological analyses

Spectroscopic and chemometric approaches to radiobiological analyses

Three dimensional collagen gels as a cell culture matrix for the study of live cells by Raman spectroscopy

Raman spectroscopy in biomedicine – non‐invasive in vitro analysis of cells and extracellular matrix components in tissues

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