Incorporation of a Disposable ESI Emitter into Inline Cartridge Extraction Mass Spectrometry Improves Throughput and Spectra Stability.

Rapid and reliable methods for detecting tumor margins are crucial for neuro-oncology. Several mass spectrometry-based methods have been recently proposed to address this problem. Inline Cartridge Extraction (ICE) demonstrates the potential for clinical application, based on ex-vivo analysis of dissected tissues, but requires time-consuming steps to avoid cross-contamination. In this work, a method of incorporating a disposable electrospray emitter into the ICE cartridge by PEEK sleeves melting is developed. It reduces total analysis time and improves throughput. The proposed setup also improves the robustness of the ICE molecular profiling as demonstrated with human glial tumor samples in that stability and reproducibility of the spectra were increased.

Mass spectrometry based proteome profiling of the exhaled breath condensate for lung cancer biomarkers search.

INTRODUCTION: Lung cancer remains the most prevalent cause of cancer mortality worldwide mainly due to insufficient availability of early screening methods for wide-scale application. Exhaled breath condensate (EBC) is currently considered as one of the promising targets for early screening and is particularly attractive due to its absolutely noninvasive collection and possibility for long-term frozen storage. EBC proteome analysis can provide valuable information about the (patho)physiological changes in the respiratory system and may help to identify in time a high risk of lung cancer. Mass spectrometry (MS) profiling of EBC proteome seems to have no alternative in obtaining the most extensive data and characteristic marker panels for screening. AREAS COVERED: This special report summarizes the data of several proteomic studies of EBC in normal and lung cancer (from 2012 to 2021, PubMed), focuses on the possible reasons for the significant discrepancy in the results, and discusses some aspects for special attention in further studies. EXPERT OPINION: The significant discrepancy in the results of various studies primarily highlights the need to create standardized protocols for the collection and preparation of EBC for proteomic analysis. The application of quantitative and targeted LC-MS/MS based approaches seems to be the most promising in further EBC proteomic studies.

Rapid estimation of tumor cell percentage in brain tissue biopsy samples using inline cartridge extraction mass spectrometry.

Tumor cell percentage (TCP) is an essential characteristic of biopsy samples that directly affects the sensitivity of molecular testing in clinical practice. Apart from clarifying diagnoses, rapid evaluation of TCP combined with various neuronavigation systems can be used to support decision making in neurosurgery. It is known that ambient mass spectrometry makes it possible to rapidly distinguish healthy from malignant tissues. In connection with this, here we demonstrate the possibility of using non-imaging ambient mass spectrometry to evaluate TCP in glial tumor tissues with a high degree of confidence. Molecular profiles of histologically annotated human glioblastoma tissue samples were obtained using the inline cartridge extraction ambient mass spectrometry approach. XGBoost regressors were trained to evaluate tumor cell percentage. Using cross-validation, it was estimated that the TCP was determined by the regressors with a precision of approximately 90% using only low-resolution data. This result demonstrates that ambient mass spectrometry provides an accurate method todetermine TCP in dissected tissues even without implementing mass spectrometry imaging. The application of such techniques offers the possibility to automate routine tissue screening and TCP evaluation to boost the throughput of pathology laboratories. Rapid estimation of tumor cell percentage during neurosurgery.

Inline cartridge extraction for rapid brain tumor tissue identification by molecular profiling.

The development of perspective diagnostic techniques in medicine requires efficient high-throughput biological sample analysis methods. Here, we present an inline cartridge extraction that facilitates the screening rate of mass spectrometry shotgun lipidomic analysis of tissue samples. We illustrate the method by its application to tumor tissue identification in neurosurgery. In perspective, this high-performance method provides new possibilities for the investigation of cancer pathogenesis and metabolic disorders.

Probabilistic model applied to ion abundances in product-ion spectra: quantitative analysis of aspartic acid isomerization in peptides.

Evaluation of post-translational modifications of protein molecules is important for both basic and applied biomedical research. Mass spectrometric quantitative studies of modifications, which do not change the mass of the protein, such as isomerization of aspartic acid, do not necessarily require the use of isotope-labelled standards. However, the accurate solution of this problem requires a deep understanding of the relationship between the mole fractions of the isomers and the peak intensities in the mass spectra. In previous studies on the isomerization of aspartic acid in short beta-amyloid fragments, it has been shown that calibration curves used for such quantitative studies often have a non-linear form. The reason for the deviation in the shape of the calibration curves from linearity has not yet been established. Here, we propose an explanation for this phenomenon based on a probabilistic model of the fragmentation process and present a general approach for the selection of fragments that can be used for quantitative studies of the degree of isomerization. Graphical Abstract.

Label-free cervicovaginal fluid proteome profiling reflects the cervix neoplastic transformation.

Cervicovaginal fluid (CVF) is a valuable source of clinical information about the female reproductive tract in both nonpregnant and pregnant women. The aim of this study is to specify the CVF proteome at different stages of cervix neoplastic transformation by label-free quantitation approach based on liquid chromatography tandem mass spectrometry (LC-MS/MS) method. The proteome composition of CVF from 40 women of reproductive age with human papillomavirus (HPV)-associated cervix neoplastic transformation (low-grade squamous intraepithelial lesion [LSIL], high-grade squamous intraepithelial lesion [HSIL], and CANCER) was investigated. Hierarchical clustering and principal component analysis (PCA) of the proteomic data obtained by a label-free quantitation approach show the distribution of the sample set between four major clusters (no intraepithelial lesion or malignancy [NILM], LSIL, HSIL and CANCER) depending on the form of cervical lesion. Multisample ANOVA with subsequent Welch's t test resulted in 117 that changed significantly across the four clinical stages, including 27 proteins significantly changed in cervical cancer. Some of them were indicated as promising biomarkers previously (ACTN4, VTN, ANXA1, CAP1, ANXA2, and MUC5B). CVF proteomic data from the discovery stage were analyzed by the partial least squares-discriminant analysis (PLS-DA) method to build a statistical model, allowing to differentiate severe dysplasia (HSIL and CANCER) from the mild/normal stage (NILM and LSIL), and receiver operating characteristic (ROC) area under the curve (AUC) were obtained on an independent set of 33 samples. The sensitivity of the model was 77%, and the specificity was 94%; AUC was equal to 0.87. CVF proteome proved to be reflect the stage of cervical epithelium neoplastic process.

Atmospheric pressure thermal ionization ion source for peptide analysis.

A novel atmospheric pressure thermal ionization (APTI) ion source was developed for the analysis of liquid samples. The feasibility of the ion source was demonstrated on peptides using two configurations-assisted by hot wire or hot surface. Microalloyed molybdenum, used as a thermal ion- emitter, notably facilitated the formation of multiply-charged ions, but fragmentation products were still observed. Peptide fragmentation accompanying thermal ionization can be used for peptide identification. The described method is promising for studies of biological samples with minimal pre-treatment.

Potassium ions are more effective than sodium ions in salt induced peptide formation.

Prebiotic peptide formation under aqueous conditions in the presence of metal ions is one of the plausible triggers of the emergence of life. The salt-induced peptide formation reaction has been suggested as being prebiotically relevant and was examined for the formation of peptides in NaCl solutions. In previous work we have argued that the first protocell could have emerged in KCl solution. Using HPLC-MS/MS analysis, we found that K(+) is more than an order of magnitude more effective in the L-glutamic acid oligomerization with 1,1'-carbonyldiimidazole in aqueous solutions than the same concentration of Na(+), which is consistent with the diffusion theory calculations. We anticipate that prebiotic peptides could have formed with K(+) as the driving force, not Na(+), as commonly believed.

Dynamically harmonized FT-ICR cell with specially shaped electrodes for compensation of inhomogeneity of the magnetic field. Computer simulations of the electric field and ion motion dynamics.

The recently introduced ion trap for FT-ICR mass spectrometers with dynamic harmonization showed the highest resolving power ever achieved both for ions with moderate masses 500-1000 Da (peptides) as well as ions with very high masses of up to 200 kDa (proteins). Such results were obtained for superconducting magnets of very high homogeneity of the magnetic field. For magnets with lower homogeneity, the time of transient duration would be smaller. In superconducting magnets used in FT-ICR mass spectrometry the inhomogeneity of the magnetic field in its axial direction prevails over the inhomogeneity in other directions and should be considered as the main factor influencing the synchronic motion of the ion cloud. The inhomogeneity leads to a dependence of the cyclotron frequency from the amplitude of axial oscillation in the potential well of the ion trap. As a consequence, ions in an ion cloud become dephased, which leads to signal attenuation and decrease in the resolving power. Ion cyclotron frequency is also affected by the radial component of the electric field. Hence, by appropriately adjusting the electric field one can compensate the inhomogeneity of the magnetic field and align the cyclotron frequency in the whole range of amplitudes of z-oscillations. A method of magnetic field inhomogeneity compensation in a dynamically harmonized FT-ICR cell is presented, based on adding of extra electrodes into the cell shaped in such a way that the averaged electric field created by these electrodes produces a counter force to the forces caused by the inhomogeneous magnetic field.

Simple synthesis of ruthenium pi complexes of aromatic amino acids and small peptides.

The interaction of [Ru(eta(6)-C(10)H(8))(Cp)](+) (Cp=C(5)H(5)) with aromatic amino acids (L-phenylalanine, L-tyrosine, L-tryptophane, D-phenylglycine, and L-threo-3-phenylserine) under visible-light irradiation gives the corresponding [Ru(eta(6)-amino acid)(Cp)](+) complexes in near-quantitative yield. The reaction proceeds in air at room temperature in water and tolerates the presence of non-aromatic amino acids (except those which are sulfur containing), monosaccharides, and nucleotides. The complex [Ru(eta(6)-C(10)H(8))(Cp)](+) was also used for selective labeling of Tyr and Phe residues of small peptides, namely, angiotensin I and II derivatives.

Initial implementation of external accumulation liquid chromatography/electrospray ionization Fourier transform ion cyclotron resonance with automated gain control.

Capillary liquid chromatography (LC) separation coupled with external accumulation Fourier transform ion cyclotron resonance (FTICR) mass spectrometry has recently been demonstrated to have significant potential for proteomics research. Accumulation of an excessive space charge external to the FTICR cell ion trap has been shown to result in increased mass measurement error, undesirable ion discrimination and/or fragmentation, potentially causing misrepresentation or incorrect assignments of lower abundance peptides in the acquired mass spectra. In this work we report on the capability of data-dependent adjustment of ion accumulation times in the course of LC separations, further referred to as automated gain control (AGC). Three different AGC approaches were evaluated based on the number of putative peptides from a tryptic digest of four casein proteins detected in the course of LC/FTICR separations. When compared with the conventional technique, AGC was found to increase, up to a factor of 3, the total number of peptides identified.