Investigation and Rapid Discrimination of Food-Related Bacteria under Stress Treatments Using IR Microspectroscopy

Because the robust and rapid determination of spoilage microorganisms is becoming increasingly important in industry, the use of IR microspectroscopy, and the establishment of robust and versatile chemometric models for data processing and classification, is gaining importance. To further improve the chemometric models, bacterial stress responses were induced, to study the effect on the IR spectra and to improve the chemometric model. Thus, in this work, nine important food-relevant microorganisms were subjected to eight stress conditions, besides the regular culturing as a reference. Spectral changes compared to normal growth conditions without stressors were found in the spectral regions of 900–1500 cm−1 and 1500–1700 cm−1. These differences might stem from changes in the protein secondary structure, exopolymer production, and concentration of nucleic acids, lipids, and polysaccharides. As a result, a model for the discrimination of the studied microorganisms at the genus, species and strain level was established, with an accuracy of 96.6%. This was achieved despite the inclusion of various stress conditions and times after incubation of the bacteria. In addition, a model was developed for each individual microorganism, to separate each stress condition or regular treatment with 100% accuracy.

The crystalline state as a dynamic system: IR microspectroscopy under electrochemical control for a [NiFe] hydrogenase

Controlled formation of catalytically-relevant states within crystals of complex metalloenzymes represents a significant challenge to structure-function studies. Here we show how electrochemical control over single crystals of [NiFe] hydrogenase 1...

Optimization of Sample Preparation Using Glass Slides for Spectral Pathology

The clinical translation of Fourier transform infrared (FT-IR) microspectroscopy in pathology will require bringing this technique as close as possible to standard practice in pathology departments. An important step is sample preparation for both FT-IR microspectroscopy and pathology. This should entail minimal disruption of standard clinical practice while achieving good quality FT-IR spectral data. In fact, the recently described possibility of obtaining FT-IR spectra of cells placed on glass substrates brings FT-IR microspectroscopy closer to a clinical application. We have now furthered this work in order to identify two different types of lung cancer cells placed on glass coverslips. Two types of sample preparation which are widely used in pathology, cytospin and smear, have been used. Samples were fixed with either methanol, used in pathology, or formalin (4% paraformaldehyde) used widely in spectroscopy. Fixation with methanol (alcohol-based fixative) removed lipids from cells causing a decrease in intensity of the peaks at 2850 cm−1 and 2920 cm−1. Nevertheless, we show for the first time that using either type of sample preparation and fixation on thin glass coverslips allowed to differentiate between two different types of lung cancer cells using either the lipid region or the fingerprint region ranging from 1800 cm−1 to 1350 cm−1. We believe that formalin-fixed cytospin samples would be preferred to study cells on thin coverslips using FT-IR microspectroscopy. This work presents a clear indication for future advances in clinical assessment of samples within pathology units to gain a deeper understanding of cells/tissues under investigation.

High-Throughput Analyses of Microplastic Samples Using Fourier Transform Infrared and Raman Spectrometry

Determining microplastics in environmental samples quickly and reliably is a challenging task. With a largely automated combination of optical particle analysis, Fourier transform infrared (FT-IR), and Raman microscopy along with spectral database search, particle sizes, particle size distributions, and the type of polymer including particle color can be determined. We present a self-developed, open-source software package for realizing a particle analysis approach with both Raman and FT-IR microspectroscopy. Our software GEPARD (Gepard Enabled PARticle Detection) allows for acquiring an optical image, then detects particles and uses this information to steer the spectroscopic measurement. This ultimately results in a multitude of possibilities for efficiently reviewing, correcting, and reporting all obtained results.

Different Approaches to FT-IR Microspectroscopy on X-ray Exposed Human Cells

Fourier-Transform Infrared microspectroscopy (μFT-IR) has been usefully applied in the analysis of the complex biological processes occurring during X-ray radiation-cell interaction. Different experimental approaches are available for FT-IR spectra collection (transmission, attenuated total reflection (ATR), and transflection modes) from cells samples. Recently, some problems have been raised about the role of transmitted and reflected components of the infrared beam in transflection mode. For this reason, we investigated two different transflection approaches for collecting spectra from cells exposed to X-ray. In the former approach, cells were grown on MirrIR slides, and for the second approach, cell pellets were prepared. In both cases, SH-SY5Y neuroblastoma cells were used. X-ray exposure was performed at doses of 2 and 4 Gy. Spectra were obtained by using both the approaches in the 600–4000 cm−1 spectral range from exposed and not-exposed samples. The main contributions from proteins, lipids, carbohydrates, and DNA were clearly evidenced in spectra obtained with the two different acquisition approaches. A comparison among them has been also reported.

Identification of a Glass Substrate to Study Cells Using Fourier Transform Infrared Spectroscopy: Are We Closer to Spectral Pathology?

The rising incidence of cancer worldwide is causing an increase in the workload in pathology departments. This, coupled with advanced analysis methodologies, supports a developing need for techniques that could identify the presence of cancer cells in cytology and tissue samples in an objective, fast, and automated way. Fourier transform infrared (FT-IR) microspectroscopy can identify cancer cells in such samples objectively. Thus, it has the potential to become another tool to help pathologists in their daily work. However, one of the main drawbacks is the use of glass substrates by pathologists. Glass absorbs IR radiation, removing important mid-IR spectral data in the fingerprint region (1800 cm−1 to 900 cm−1). In this work, we hypothesized that, using glass coverslips of differing compositions, some regions within the fingerprint area could still be analyzed. We studied three different types of cells (peripheral blood mononuclear cells, a leukemia cell line, and a lung cancer cell line) and lymph node tissue placed on four different types of glass coverslips. The data presented here show that depending of the type of glass substrate used, information within the fingerprint region down to 1350 cm−1 can be obtained. Furthermore, using principal component analysis, separation between the different cell lines was possible using both the lipid region and the fingerprint region between 1800 cm−1 and 1350 cm−1. This work represents a further step towards the application of FT-IR microspectroscopy in histopathology departments.

Synchrotron IR-microspectroscopy-based visualization of molecular and chemical interactions between dental cement, biomimetic composite and native dental tissue

The low affinity of composite materials for the hard tissue of human teeth poses a challenge to restorative dentists. This study was undertaken to explore molecular and chemical characteristics of the interface between the dental cement, the buffer layer formed from a next generation biomimetic material that mimics the organic mineral composition of human enamel and dentin, and the intact native hard dental tissue. Seven plane-parallel dental slices were analyzed using synchrotron IR microspectroscopy. The obtained absorption spectra of functional molecular groups were organized into cluster maps. This allowed us to identify the intact tissue, the adhesive agent and the biomimetic layer at their interface and to localize and measure concentrations of functional groups involved in the integration of the biomimetic composite into the hard tissue of the human tooth. The proposed biomimetic material is based on nanocrystal carbonate-substituted calcium hydroxyapatite synthesized from a biogenic calcium source and a complex of basic polar amino acids copying the composition of the human tooth and can form a functional bond with hard dental tissue.