Klinisch-spektroskopische Diagnostik

Arbeitsgruppe Prof. Dr. Ute Neugebauer

Ute Neugebauer, Univ.-Prof. Dr.

Arbeitsgruppenleiterin
Albert-Einstein-Straße 9
07745 Jena Google Maps – LageplanExterner Link

Mitarbeiter der AG Neugebauer

  1. Neugebauer, Ute, Univ.-Prof. Dr.

    Professur Klinisch-Spektroskopische Diagnostik
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44 Publikationen filtern

Die Publikationen filtern

  1. A comparative study of robustness to noise and interpretability in U-Net-based denoising of Raman spectra

    ErscheinungsjahrErschienen in:Spectrochimica Acta - Part A: Molecular and Biomolecular Spectroscopy A. Mokari, S. Eiserloh, O. Ryabchykov, U. Neugebauer, T. Bocklitz
    Universitätsbibliographie Jena:
    fsu_mods_00026011Externer Link

  2. Insights into chlamydial infection at the sub-cellular level using label-free Raman spectroscopy in comparison to electron microscopy

    ErscheinungsjahrErschienen in:Journal of Biological Chemistry N. Unger, E. Liebler-Tenorio, R. Guliev, S. Eiserloh, S. Nietzsche, F. Nowak, S. Zuchantke, C. Berens, C. Schnee, U. Neugebauer
    Intracellular infections are difficult to study as the host cell protects the pathogen from direct observation from the outside. Transmission electron microscopy (TEM) is the most commonly used method for subcellular analysis. However, sample preparation is based on fixation which prevents continuous observation. Here, we focus on the obligate intracellular bacterium Chlamydia abortus. It causes infections primarily in small ruminant livestock, and can also be transmitted to humans, where it can cause disease. Diagnosis is difficult, requiring PCR or cell culture. At the moment, non-invasive methods for the direct study of intracellular infections are rare and not yet established in routine analysis. In this study, we present 3D confocal Raman imaging as a non-invasive tool to investigate and characterize the infection directly inside intact host cells without the need for any purification step and compare the results to established conventional TEM. A 2D cell culture infection model with Buffalo Green Monkey kidney cells was employed and infection with Chlamydia abortus S26/3 was characterized at different time points post infection. Using multivariate statistical data analysis, high-quality false color image stacks were generated from the Raman data. Two Chlamydia morphoforms, elementary body and reticulate body, could be distinguished based on their Raman spectral features: reticulate bodies are characterized by prominent lipid and nucleic acid signals while elementary bodies revealed higher carbohydrate and protein signals. This provides complementary information to TEM analysis where morphoforms are differentiated based on size and contrast. The complementary nature of both imaging methods is discussed in the manuscript.
    Universitätsbibliographie Jena:
    fsu_mods_00029339Externer Link

  3. Molecular Insights into the Heme-Binding Potential of Plant NCR247-Derived Peptides

    ErscheinungsjahrErschienen in:ChemBioChem : a European journal of chemical biology S. Vaidya, D. Rathod, A. Ramoji, U. Neugebauer, D. Imhof
    Heme is involved in many critical processes in pathogenic bacteria as iron acquisition by these microorganisms is achieved by either direct uptake of heme or use of heme-binding proteins called hemophores. Exploring the underlying mechanisms on a molecular level can open new avenues in understanding the host-pathogen interactions. Any imbalance of the heme concentration has a direct impact on the bacterial growth and survival. Thus, heme-regulated proteins that are involved in heme homeostasis poise to be promising targets for research. Similarly, naturally occurring compounds, including cysteine-rich peptides from either plant secondary metabolites or venom toxins from vertebrates and invertebrates, have been studied for their therapeutic potential. NCR247 is such a cysteine-rich peptide, known to be crucial for nitrogenase activity in M. truncatula and its symbiotic relation with S. meliloti. NCR247-derived peptides were suggested to serve as high-affinity heme-binding molecules with remarkable heme-capturing properties. A comprehensive biochemical and computational analysis of NCR247-derived peptides, however, redefines their heme-binding capacity and consequently their potential therapeutic role.
    Universitätsbibliographie Jena:
    fsu_mods_00019376Externer Link

  4. Super-resolution fluorescence MINFLUX microscopy to study lipid dynamics in live-cell plasma membranes

    Erscheinungsjahr A. Koerfer
    The Plasma Membrane (PM) of eukaryotic cells separates the intracellular environment from the extracellular space and plays a crucial role in regulating material exchange, signal transduction, and host-pathogen interactions. It consists of a heterogeneous lipid bilayer with embedded proteins, enabling localised processes such as endocytosis, exocytosis, and viral assembly. However, its small size and rapid dynamics make direct study challenging. A prominent example is the assembly and release of Human Immunodeficiency Virus type 1 (HIV-1). During viral assembly, the viral protein Gag binds the cytosolic leaflet of the PM, triggering clustering and membrane curvature that leads to budding of new virions (~120 nm). This is accompanied by local enrichment of cholesterol and saturated lipids, though the underlying mechanisms remain unclear. This thesis investigates lipid dynamics in the live-cell PM using the super-resolution technique Minimal Photon Flux (MINFLUX) microscopy. MINFLUX enables tracking of individual fluorescent lipid analogues with high spatial (nanometre) and temporal resolution. Both 2D and 3D MINFLUX tracking are used to study lipid diffusion and enrichment during the formation of HIV Virus-Like Particles (VLPs) induced by Gag. 3D MINFLUX reveals membrane topography at ~20 nm resolution, allowing classification of intermediate budding stages. Fully formed VLPs measure 130 ± 5 nm, consistent with literature. Correlating structural features with lipid enrichment and diffusion provides insights not accessible with 2D data alone. Additionally, a custom analysis workflow integrating confocal imaging MINFLUX tracking is presented, enabling quantitative correlation between lipid dynamics and Gag localisation. Overall, this work establishes an optimised MINFLUX-based workflow for studying lipid dynamics in live-cell PMs and demonstrates the importance of integrating structural information to understand membrane remodelling during viral budding.
    Universitätsbibliographie Jena:
    fsu_mods_00029829Externer Link

  6. Qualitative comparison of decalcifiers for mouse bone cryosections for subsequent biophotonic analysis

    ErscheinungsjahrErschienen in:Scientific Reports S. Mandal, R. Motganhalli Ravikumar, A. Tannert, A. Urbanek, R. Guliev, M. Naumann, S. Coldewey, U. Dahmen, L. Carvalho, L. Bastião Silva, U. Neugebauer
    Bone tissue, with its complex structure, often necessitates decalcification of the hard tissue for ex vivo morphological studies. The choice of a suitable decalcification method plays a crucial role in preserving desired features and ensuring compatibility with diverse imaging techniques. The search for a universal decalcification method that is suitable for a range of biophotonic analyses remains an ongoing challenge. In this study, we systematically assessed five standard bone decalcification protocols, encompassing strong mineralic acids (3% and 5% nitric acid), a commercially available formulation of hydrochloric and formic acid), as well as weak organic acids (5% trichloroacetic acid and 8% formic acid), and a chelating agent (25% ethylenediamine-tetraacetic acid) with varying decalcification durations, using mouse long bones as our experimental model. Our imaging analysis panel included classical histological staining (Hematoxylin and Eosin, H&E), immunofluorescence staining, and label-free Raman microspectroscopic imaging. We used cryosections instead of paraffin sections since paraffin interferes with tissue Raman signals. This approach is not as commonly used as it is more prone to handling artifacts, but is the preferred method for subsequent Raman analysis. Decalcification efficacy was evaluated based on various qualitative and some quantitative imaging parameters by 2–3 independent observers. Our systematic approach revealed that the chelating agent, when used for 24 h, optimally preserved bone features and, thus, would be the ideal decalcifying agent for comprehensive subsequent analysis. However, the choice of decalcifier and the ideal decalcification duration may vary depending on the type and thickness of bone, necessitating tailored adjustments to meet specific experimental requirements.
    Universitätsbibliographie Jena:
    fsu_mods_00019104Externer Link

  7. Investigation of Collagen Crosslinks Introduced By a Femtosecond Laser

    ErscheinungsjahrErschienen in:2025 Conference on Lasers and Electro-Optics Europe & European Quantum Electronics Conference (CLEO/Europe-EQEC) D. Fischer, A. Stöcker, A. Tannert, T. Koch, U. Neugebauer, R. Ackermann, J. Missbach-Guentner, S. Nolte, C. Rußmann
    Universitätsbibliographie Jena:
    fsu_mods_00027637Externer Link

  8. Characterizing Metabolic Shifts in Septic Murine Kidney Tissue Using 2P-FLIM for Early Sepsis Detection

    ErscheinungsjahrErschienen in:Bioengineering S. Greiner, M. Ebrahimi, M. Rodewald, A. Urbanek, T. Meyer-Zedler, M. Schmitt, U. Neugebauer, J. Popp
    In this study, thin mouse kidney sections from healthy mice and those infected leading to acute and chronic sepsis were examined with two-photon excited fluorescence lifetime imaging (2P-FLIM) using the endogenous fluorescent coenzymes nicotinamide adenine dinucleotide (NADH) and flavin adenine dinucleotide (FAD). The results presented show that this approach is a powerful tool for investigating cell metabolism in thin tissue sections. An adapted measurement routine was established for these samples by performing a spectral scan, identifying a combination of two excitation wavelengths and two detection ranges suitable for detailed scan images of NADH and FAD. Selected positions in thin slices of the renal cortex of nine mice (three healthy, three with chronic sepsis, and three with acute sepsis) were studied using 2P-FLIM. In addition, overview images were obtained using two-photon excited fluorescence (2PEF) intensity. This study shows that healthy kidney slices differ considerably from those with acute sepsis with regard to their fluorescence lifetime signatures. The latter shows a difference in metabolism between the inner and outer cortex, indicating that outer cortical tubular cells switch their metabolism from oxidative phosphorylation to glycolysis in kidneys from mice with acute sepsis and back in later stages, as seen for mice with chronic infections. These findings suggest that 2P-FLIM could serve as a powerful tool for early-stage sepsis diagnosis and monitoring metabolic recovery during treatment.
    Universitätsbibliographie Jena:
    fsu_mods_00021975Externer Link

  9. Long-term device stability for Raman spectroscopy

    ErscheinungsjahrErschienen in:The analyst: the analytical journal of the Royal Society of Chemistry S. Guo, A. Ramoji, A. Pistiki, H. Yilmaz, U. Glaser, D. Vasquez-Pinzon, I. Schie, U. Neugebauer, A. Silge, J. Popp, T. Bocklitz
    Universitätsbibliographie Jena:
    fsu_mods_00025267Externer Link

  10. Nanochemical Cell-Surface Evaluation in Photothermal Spectroscopic Imaging of Antimicrobial Interactions in the Model System Bacillus subtilis and Vancomycin

    ErscheinungsjahrErschienen in:Analytical chemistry M. Ali, R. Schneider, A. Strecker, N. Krishnakumar, S. Unger, M. Soltaninezhad, J. Kirchhoff, A. Tannert, K. Dragounova, R. Heintzmann, A. Müller, C. Krafft, U. Neugebauer, D. Täuber
    The power of photothermal spectroscopic imaging to visualize antimicrobial interactions on the surfaces of individual bacteria cells has been demonstrated on the model system Bacillus subtilis and vancomycin using mid-infrared photoinduced force microscopy (PiF-IR, also mid-IR PiFM). High-resolution PiF contrasts obtained by merging subsequent PiF-IR scans at two different illumination frequencies revealed chemical details of cell wall destruction after 30 and 60 min incubation with vancomycin with a spatial resolution of ∼5 nm. This approach compensates for local intensity variations induced by near-field coupling of the illuminating electric field with nanostructured surfaces, which appear in single-frequency contrasts in photothermal imaging methods, as shown by Anindo et al. [J. Phys. Chem. C 2025, 129, 4517. DOI: 10.1021/acs.jpcc.4c08305]. Known spectral shifts associated with hydrogen bond formation between vancomycin and the N-acyl-d-Ala₄ -d-Ala₅ termini in the peptidoglycan cell wall have been observed in chemometrics of PiF-IR spectra from treated and untreated B. subtilis harvested after 30 min from the same experiment. Spectral signatures of the vancomyin interaction have been located in the piecrust of a progressing septum with ∼10 nm resolution using PiF contrasts of three selected bands of a PiF-IR hyperspectral scan of an individual B. subtilis cell harvested after 30 min incubation. Our results are complemented by a discussion of imaging artifacts and the influence of parameter settings supporting further development toward standardization in the application of PiF-IR for visualizing the chemical interaction of antibiotics on the surface of microbes with few nanometer resolution.
    Universitätsbibliographie Jena:
    fsu_mods_00029357Externer Link

  11. Blood biomarker and Raman analysis of leukocytes in the multicenter INTELLIGENCE trials: at the Intensive Care Unit (INTELLIGENCE-1) and the Emergency Department (INTELLIGENCE-2)

    ErscheinungsjahrErschienen in:Shock: molecular, cellular and systemic pathobiological aspects and therapeutic approaches E. Giamarellos-Bourboulis, D. Thomas-Rüddel, A. Pistiki, A. Ramoji, O. Ryabchykov, K. Azam, K. Toutouzas, A. Prekates, S. Karatzas, C. Mathas, A. Kotsaki, S. Chalvatzis, N. Antonakos, G. Damoraki, J. Rüger, F. Knorr, N. Arend, A. Silge, I. Schie, J. Eugen-Olsen, T. Bocklitz, M. Bauer, J. Winning, M. Kiehntopf, J. Popp, F. Bloos, U. Neugebauer
    Background: – Sepsis, defined as life-threatening organ dysfunction caused by a dysregulated host response to infection, goes along with a complex and not yet fully understood host response. Despite many different biomarkers, optimal screening tools to identify patients with sepsis are still needed. In a previous, single-center clinical trial with well-defined and well-separated patient cohorts, a new biophotonic marker based on the Raman spectroscopic characterization of leukocytes showed added value to stratify patients and identify infection and sepsis. Results: – In the INTELLIGENCE studies, 279 patients from six centres in two countries were analysed and the Raman spectra of ~1500 leukocytes per patient measured within only 1 hour. This marks a huge step from bench to bedside regarding usability and technology readiness level of Raman spectroscopy in multicentre clinical trials. With a discriminatory power comparable to individual conventional biomarkers (CRP, PCT, IL-6, suPAR), the Raman score of leukocytes could not provide added value to the clinical discrimination of sepsis in the INTELLIGENCE-1 study cohorts of intensive care patients with infections. When translating the classification model from INTELLIGENCE-1 to the heterogeneous patient cohort recruited at the emergency department of the double-blinded INTELLIGENCE-2 trial, the Raman score failed to provide added value. Conclusions: – In theory, Raman assessment of leukocytes might still be a promising tool for sepsis diagnosis and patient stratification, but there is more basic, translational and clinical research needed to refine its usability and clinical role, probably also taking questions of the pathophysiology of the dysregulated host response for a phenotype stratification into account.
    Universitätsbibliographie Jena:
    fsu_mods_00029323Externer Link

  12. Raman Spectroscopy Can Identify Acute and Persistent Biochemical Changes in Leukocytes From Patients With COVID-19 and Non-COVID-19-Associated Sepsis

    ErscheinungsjahrErschienen in:Biotechnology journal A. Ramoji, P. Baumbach, O. Ryabchykov, A. Pistiki, J. Rueger, D. Pinzon, A. Silge, S. Deinhardt-Emmer, I. Schie, K. Weber, C. Neu, U. Neugebauer, M. Kiehntopf, T. Bocklitz, J. Popp, S. Coldewey
    Sepsis remains a major clinical challenge, often resulting in long-term physiological and immunological disturbances. This study employed high-throughput single-cell Raman spectroscopy to analyze the biochemical profiles of peripheral blood leukocytes from patients with non-COVID-19 and COVID-19-associated sepsis. Leukocytes were assessed at multiple timepoints, including the acute phase (Days 3 and 7 after sepsis onset) and late recovery phase (6 and 12 months after sepsis onset). Raman spectroscopic profiles of leukocytes showed clear separation between healthy controls and sepsis patients during the acute phase with high balanced accuracy (BAcc: 95%–98%). Spectral differences between acute and recovery phases (BAcc: 84%–97%) and between recovery-phase leukocytes and those from healthy controls (BAcc: 81%–90%) were also observed, indicating long-lasting molecular alterations. Furthermore, distinct profiles were identified between non-COVID-19 and COVID-19-associated sepsis during the acute phase (BAcc: 65%–71%) and in the late-recovery phase (BAcc: 71%–83%). These findings demonstrate that Raman spectroscopy enables label-free, high-throughput profiling of leukocyte biochemistry across the sepsis trajectory. This suggests that Raman spectroscopy is a promising tool for high-throughput screening, offering insights into the biomolecular changes in sepsis and providing a diagnostic platform to differentiate between sepsis etiologies, a significant advancement in the field of sepsis diagnostics.
    Universitätsbibliographie Jena:
    fsu_mods_00027266Externer Link
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