Origanum L. (Lamiaceae) is an important genus of medicinal and aromatic plants used in traditional medicine since ancient times as culinary herbs and remedies. The aim of the present study was to evaluate the chemical composition, as well as the biochemical and cellular activities of freshly prepared Origanum majorana L. essential oil (OmEO) in an Alzheimer’s disease (AD) amyloid beta1-42 (Aβ1-42) rat model. OmEO (1% and 3%) was inhaled for 21 consecutive days, while Aβ1-42 was administered intracerebroventricularly to induce AD-like symptoms. Our data demonstrate that OmEO increased antioxidant activity and enhanced brain-derived neurotrophic factor (BDNF) expression, which in concert contributed to the improvement of cognitive function of animals. Moreover, OmEO presented beneficial effects on memory performance in Y-maze and radial arm-maze tests in the Aβ1-42 rat AD model.

Fetal growth restriction (FGR) affects about 3% to 8% of pregnancies, leading to higher perinatal mortality and morbidity. Current strategies for detecting fetal growth impairment are based on ultrasound inspections. However, antenatal detection rates are insufficient and critical in countries with substandard care. To overcome difficulties with detection and to better discriminate between high risk FGR and low risk small for gestational age (SGA) fetuses, we investigated the suitability of risk assessment based on the analysis of a recently developed proteome profile derived from maternal serum in different study groups. Maternal serum, collected at around 31 weeks of gestation, was analyzed in 30 FGR, 15 SGA, and 30 control (CTRL) pregnant women who delivered between 31 and 40 weeks of gestation. From the 75 pregnant women of this study, 2 were excluded because of deficient raw data and 2 patients could not be grouped due to indeterminate results. Consistency between proteome profile and sonography results was obtained for 59 patients (26 true positive and 33 true negative). Of the proteome profiling 12 contrarious grouped individuals, 3 were false negative and 9 were false positive cases with respect to ultrasound data. Both true positive and false positive grouping transfer the respective patients to closer surveillance and thorough pregnancy management. Accuracy of the test is considered high with an area-under-curve value of 0.88 in receiver-operator-characteristics analysis. Proteome profiling by affinity-mass spectrometry during pregnancy provides a reliable method for risk assessment of impaired development in fetuses and consumes just minute volumes of maternal peripheral blood. In addition to clinical testing proteome profiling by affinity-mass spectrometry may improve risk assessment, referring pregnant women to specialists early, thereby improving perinatal outcomes.

Precision medicine requests accurate serological inspections to precisely stratify patients for targeted treatment. Intact transition epitope mapping analysis proved surrogate seroconversion of a model organism’s serum when spiked with a monoclonal murine anti-Ovalbumin antibody (mAb) with epitope resolution. Isolation of the IgG fraction from blood serum applied two consecutive protein precipitation steps followed by ultrafiltration and resulted in an ESI-MS analysis-ready IgG preparation. For epitope mapping by epitope extraction, the Ovalbumin antigen was digested with trypsin. After desalting, the peptide mixture was added to the ESI-MS-ready IgG preparation from mAb-spiked serum and the solution was incubated to form an immune complex between the Ovalbumin-derived epitope peptide and the anti-Ovalbumin mAb. Then, the entire mixture of proteins and peptides was directly electrosprayed. Sorting of ions in the mass spectrometer’s gas phase, dissociation of the immune complex ions by collision-induced dissociation, and recording of the epitope peptide ion that had been released from the immune complex proved the presence of the anti-Ovalbumin mAb in serum. Mass determination of the complex-released epitope peptide ion with isotope resolution is highly accurate, guaranteeing high specificity of this novel analysis approach, which is termed Intact Transition Epitope Mapping—Serological Inspections by Epitope EXtraction (ITEM—SIX).

Immunoassays for cardiac troponin, such as the Elecsys^® hs-TnT, have become the gold standard for myocardial infarction diagnostics. While various protein/chemical factors affecting the troponin complex and thus its diagnostic accuracy have been investigated, the role of coding single-nucleotide polymorphisms remains underexplored. To evaluate potential cSNP-induced interference with antibody binding in the Elecsys^® hs-TnT immunoassay, we applied ITEM-FOUR, a mass spectrometry-based method that quantifies changes in antibody binding upon amino acid substitutions in epitope peptides. Candidate cSNPs were selected from the dbSNP database and were mapped to human cardiac troponin T by molecular modeling. Consuming micromolar antibody concentrations and microliter sample volumes, two wild-type and 17 cSNP-derived variant epitope peptides—six for monoclonal antibody M7 and eleven for monoclonal antibody M11.7—were investigated to reveal the binding motifs “V₁₃₁-K₁₃₄-E₁₃₈-A₁₄₂” for M7 and “E₁₄₆-I₁₅₀-R₁₅₄-E₁₅₇” for M11.7. Loss of binding to M11.7 was observed for substitutions Q148R (rs730880232), R154W (rs483352832), and R154Q (rs745632066), whereas the E138K (rs730881100) exchange disrupted binding of M7. Except for cSNP Q148R, they are associated with cardiomyopathies, placing affected individuals at risk of both underlying heart disease and false-negative hs-TnT assay results in cases of myocardial infarction. Our results highlight the need to account for cSNP-related interferences in antibody-based diagnostics. ITEM-FOUR offers a powerful approach for tackling this challenge, fostering next-generation assay development.

Studying protein structure dynamics is key to understanding protein function modulation. Alternative protein conformations are well discriminated from each other by nanoESI mass spectrometry. The shapeshifting IgG antibodies and protein G are present as compactly folded native conformations in neutral buffered solution, identified by molecular ions with narrow charge state distributions, relatively few charges, and small collisional cross sections. Simultaneously present extended but nevertheless native conformations produced additional ions with higher charge states, different charge state distributions, and larger collisional cross sections. Computed collisional cross sections from “o-shape” and extended “I-shape” protein G 3D structures match to experimental data, indicating equilibrium, and an “o2I” flip process. Likewise, “m-shape” and “Y-shape” IgGs are two of reversibly adopted antibody conformations which interchange by an “m2Y” flip. Burying Fc receptor binding sites by adopting the m-shape prevents antibody-based initiation of humoral and cellular immune system responses prior to antigen contact. The “m2Y” flip provides robust regulation of opsonophagocytosis, an important immune system mechanism.

Studying protein structure dynamics is key to understanding protein function modulation. Alternative protein conformations are well discriminated from each other by nanoESI mass spectrometry and ion mobility measurements. Experimentally determined collisional cross-sections were compared to calculated collisional cross-sections of fifteen peptides, single-domain proteins, and protein complexes. The multi-domain proteins investigated here, four immunoglobulin G (IgG) antibodies and protein G, are present as compacted/folded “native” conformations in neutral buffered solutions, and they are identified by molecular ions with narrow charge-state distributions, relatively few charges, and small collisional cross-sections. Simultaneously present extended/folded but nevertheless “native” conformations produced additional ions with higher charge states, different charge-state distributions, and larger collisional cross-sections. Computed collisional cross-sections from compacted “o-shape” and extended “l-shape” protein G three-dimensional (3D) structures match experimental data, indicating equilibrium, and suggest a dynamic “o2l” flip process. Likewise, “m-shape” (compacted) and “Y-shape” (extended) IgGs are regarded as two supposedly reversibly adopted antibody conformations which may interchange by an “m2Y” flip. Adopting an m-shape would prevent an antibody-based initiation of humoral and cellular immune system responses, such as opsonophagocytosis, prior to antigen contact, which stands in line with the rearrangement hypothesis.