The aim of this study was to examine the hepatoprotective activity of multicomponent mixtures of natural origin in the BALB/C mouse model (n = 59), with acute direct toxic liver injury (DTLI) induced by the administration of streptozotocin (STZ) (100 mg/kg) in combination with a high-fat and high-fructose diet (HFFD). The hepatoprotective activity of activated hydrolytic lignin (Bp-Cx-1), methanolic fraction of Bp-Cx-1 (Bp-Cx-M) and isoflavones from kudzu Pueraria lobata roots (IFL) were evaluated on molecular level using mass spectrometry (MS)-based omics technologies. Untargeted label-free DIA quantitation resulted in 7214 protein groups identification (FDR 1%) after filtering across 40 liver tissue extracts. All treatment groups were closer to the control samples on the liver proteomic landscape compared to the untreated DTLI group, with the best results shown for the Bp-Cx-M and IFL groups. In order to identify differences between specific groups, we applied the post hoc Dunn's test and used Hedges' g as the effect size metric, revealing 64 proteins that tended to return to their normal level after treatment. In-depth proteomic liver tissue analysis enabled us not only to reveal the main pathways such as inflammation and oxidative stress, which are in a good agreement with DTLI and non-alcoholic liver disease pathophysiology, but also to evaluate hepatoprotective activity of multicomponent mixtures of natural origin containing polyphenols and mostly associated with protein metabolism (e.g., PSMD7, HCFC1) and deubiquitination pathways (e.g., UCHL3). It is worth noting that the Bp-Cx-1 isolated methanol fraction (Bp-Cx-M) demonstrated a pronounced increased hepatoprotective activity compared to the parent material due to the enrichment with active components such as polyphenols. Consistent with the proteomic findings of restored ubiquitin-proteasome function, assessment by comet assay revealed that treatments with Bp-Cx-M and IFL significantly reduced DNA damage by 50% compared to the untreated DTLI group. The developed MS-based multi-omics approach may be implemented for the robust and high-throughput screening method during assessment of new hepatoprotective agents of synthetic or natural origin.

For successful exploration of near space, construction of planetary bases and exploration of the planets of the solar system, it is necessary to fill the gaps in understanding the molecular mechanisms of the human body’s response to space flight (SF) conditions. Subsequently, this will make it possible to get closer to the discovery of potential molecular targets for protection against adverse processes occurring in the body under the influence of SF conditions. The goal of the work was to assess the effect of a 6-month space flight on the proteome of dried blood spots of cosmonauts. As a result of proteomic analysis, processes with highly and moderately enriched genes were identified. Among them, processes with reliable dynamics were identified on the 7th day of SF, as well as on the 3rd and 6th months of flight: these are clusters of energy processes, presentation of molecules on the membrane, initiation of immune defense, proteostasis, and metabolism. Biological processes were identified in which the representation of proteins decreased most significantly, which was reflected in a weakening of activity in the presentation of molecules on the membrane, the initiation of immune defense, as well as in the mechanisms of proteostasis in the acute period of adaptation to the factors of the initial stage of flight. It has been shown that there is a close interaction with proteins of the cytoskeleton organization of proteins that disappear or reappear in the dried blood spots of proteome during flight, and they relate to processes whose activity significantly decreased during SF (immune system, proteostasis, metabolism).

Pancreatic cancer (PC) is the sixth leading cause of cancer-related deaths worldwide. Patients with pancreatic ductal adenocarcinoma (PDAC), the most common type of PC, have a 5-year survival rate of approximately 10%. This low survival rate is mainly attributed to late-stage diagnoses and the lack of robust screening methods. Several serum proteins have been proposed as potential PDAC biomarkers, but they have not been introduced into clinical practice due to their low sensitivity and specificity. Therefore, the identification of new PDAC biomarkers remains highly important, and multiple reaction monitoring (MRM), a highly accurate mass spectrometry (MS) technique, can be used for this purpose. Using MRM MS analysis, we estimated the concentrations of 103 proteins in peripheral blood plasma from 132 participants: patients with newly diagnosed PDAC at different stages and healthy individuals. We identified six proteins that were differentially presented between healthy controls and patients with PDAC at all stages (adjusted p-value < 0.01), and that were associated with survival rates for 23 months. A developed cross-validated model based on these six proteins showed an average accuracy of 90% in distinguishing between early-stage PDAC and healthy controls (AUC = 0.933). However, further research is needed to implement this model in clinical practice.

Advances in liquid chromatography/mass spectrometry (LC-MS) have enabled proteome-wide quantitation in minutes, achieving rate of 1000 analyses per day. This necessitates revisiting the rapid sample preparation approaches to match this data acquisition speed. Despite the fact that these approaches have been developed decades ago, their application in quantitative ultrafast proteomics and comprehensive comparison of their performance under different conditions have not been explored. In this study, the ultrasound, microwave irradiation, and elevated temperature-assisted approaches for accelerated protein reduction, alkylation, and trypsin digestion were compared. Validation was carried out with label-free quantitative LC-MS/MS and fragmentation-free DirectMS1 methods of shotgun proteome analyses of Saccharomyces cerevisiae, human cell lines, and winter wheat shoots. These data acquisition methods were applied in ultrafast implementations employing 5 to 16 min LC gradients. Human-yeast proteome mixtures were used as standards to evaluate quantitation accuracy of the sample preparation workflows. Our findings indicate that the reduced time of sample preparation insignificantly decreased efficiency of reduction, alkylation, and digestion, yet, preserved reproducible peptide and protein identification. We also found that the 30-min microwave-assisted and overnight trypsin digestion yielded comparable quantitation accuracy in ultrafast analyses using DirectMS1 method.

Early recognition of a risk of Alzheimer's disease (AD) remains a global challenge, and blood proteomic markers are of particular interest for wide-scale diagnostic use. Quantitative multiple reaction monitoring (MRM) approach demonstrates good reproducibility in the characteristic changes in the levels of reported candidate biomarkers (CBs) in different cohorts in AD. Following up on our previous study, we performed a joint analysis of 331 blood plasma samples from two different clinical cohorts of participants, comprising a total of 95 samples from patients with AD, 136 samples from patients with mild cognitive impairment (MCI), and 100 samples from controls. The obtained results confirm the significance of 37 CBs. A logistic regression-based algorithm was used to build protein classifiers, and a total of 21 important proteins were selected, 13 of which (ORM1, APOA4, LBP, HP, FN1, BCHE, APOE, PZP, A1BG, TF, SERPINA7, TTR, and F12) formed a universal panel that demonstrated strong classification performance in distinguishing AD patients from controls (ROC-AUC = 0.90) and in separating stable and progressing patients with MCI (ROC-AUC = 0.81). Overall, the analysis confirms the high potential of the MRM method for validating CBs in independent cohorts.

INTRODUCTION:Identifying early risks of developing Alzheimer's disease (AD) is a major challenge as the number of patients with AD steadily increases and requires innovative solutions. Current molecular diagnostic modalities, such as cerebrospinal fluid (CSF) testing and positron emission tomography (PET) imaging, exhibit limitations in their applicability for large-scale screening. In recent years, there has been a marked shift toward the development of blood plasma-based diagnostic tests, which offer a more accessible and clinically viable alternative for widespread use. Furthermore, advances in large-scale proteomics technologies have boosted an interest in identifying novel biomarkers and developing panels of AD-associated proteins. AREAS COVERED:This review mainly examines the results of recent searches for proteomic markers of AD in blood plasma (from 2022-2024 PubMed), focuses on some aspects for special attention in further studies, and discusses the prospects for their further application. EXPERT OPINION:Recent advances in AD plasma/serum proteomic studies are largely driven using novel Olink/PEA and SomaScan/aptamer technologies, which complement the 'gold standard' of MS-based quantitative proteomics (MRM/SRM), and particularly expand the capabilities for studying low-abundant proteins.

AIM:To identify the features of plasma, platelet hemostasis, and proteomic composition of the blood plasma in patients with acute myocardial infarction (AMI) and healthy volunteers after COVID-19. MATERIAL AND METHODS:The study included patients with AMI who have recently had COVID-19 (AMI-post-COVID, n=56) and patients with AMI who have not recently had COVID-19 (AMI-control, n=141). Healthy volunteers constituted the control groups and were also divided into control-post-COVID (n=32) and control-control (n=71) groups. Previous SARS-CoV-2 infection was determined by anti-N IgG in the blood serum, the level of which persists for 6-10 months after the disease. Hemostasis was evaluated by thromboelastometry (on whole blood), thrombodynamics (on platelet-poor plasma), fibrinolysis, impedance aggregometry, and proteomic analysis. RESULTS:The AMI-post-COVID and AMI-control groups had higher values of thrombus growth rate, size and density based on the data of thromboelastometry and thrombodynamics, as well as increased concentrations of the complement system components, proteins regulating the state of the endothelium, and a number of acute-phase and procoagulant proteins compared to the control groups. Furthermore, in the AMI-post-COVID group, compared to the AMI-control group, the thrombus density was lower, and its lysis rates were higher when measured by the thrombodynamics method on platelet-poor plasma, while the platelet aggregation induced by ADP and thrombin was higher. Also, in the control-post-COVID group, compared to the control-control group, the thrombus formation rate was lower, whereas, in contrast, the thrombus size as measured by the thrombodynamics method and the platelet aggregation induced by arachidonic acid and thrombin were higher. In addition, in the AMI-post-COVID group, compared to the AMI-control group, the concentrations of proteins involved in inflammation and hemostasis were lower. CONCLUSION:Patients with AMI who have recently had COVID-19 are characterized by a less pronounced activation of the immune response compared to patients with AMI who have not had COVID-19. This may be due to long-term chronic inflammation and depletion of components of the immune activation system after SARS-CoV-2 infection. Long-term activation of the hemostasis system in both patients with AMI and healthy volunteers after COVID-19 is primarily due to the platelet component of hemostasis.

Aim. To study the plasma proteome of patients with type 1 acute myocardial infarction (AMI) to identify potential markers for long-term prognosis of the risk for developing cardiovascular complications.Material and methods. The study included 64 patients with type 1 AMI with and without ST segment elevation who underwent primary percutaneous coronary intervention upon admission. The following information on cardiovascular events was collected for 36 months after admission: death from cardiovascular pathology, recurrent AMI, stroke, repeat myocardial revascularization and/or endarterectomy. Peripheral blood sampling followed by a plasma proteome analysis using chromatography-mass spectrometry was performed in all patients before hospitalization.Results. During 36 months after hospitalization, cardiovascular complications were detected in 23 (36%) patients. These patients were included in the group with an unfavorable prognosis, while the remaining patients made up the group with a positive prognosis. A mass spectrometric analysis of the plasma proteome and comparison of the groups identified seven differentially represented proteins. Also, a multivariate regression analysis, ROC curves, and Kaplan-Meier models showed that four proteins (apolipoprotein C1, complement factor H, di-N-acetylchitobiase, and ficolin-2) were predictors of the risk for developing cardiovascular complications in the long term. An integrated parameter was developed that took into account the plasma concentrations of all four above proteins. This parameter was used to construct a model for assessing the risks of unfavorable long-term prognosis in AMI patients with a sensitivity of 87% and a specificity of 78%.Conclusion. The study results demonstrated that plasma concentrations of apolipoprotein C1, complement factor H, di-N-acetylchitobiase, and ficolin-2 are reliable prognostic markers for assessing the risks of cardiovascular events in patients with AMI in the long term.

Chromatography-mass spectrometry was used to analyze dried blood spots collected from 6 female volunteers in a 3-d dry immersion (DI) in order to detect proteins involved in acute adaptation of the female physiological systems. Sampling was performed before DI, every day in DI and on the next day after DI completion. Proteomic analysis evidenced an immediate reaction of proteins involved in carbohydrate metabolism and proteasome degradation of proteins. On DI day-2, we saw a reaction of immune response proteins and decreased PEDF level, an angiogenesis inhibitor that controls many processes, including heart protection from injuries. On DI-3, the response was joined by proteins participating in processes of the complement cascade and coagulation, platelet degranulation, hemostasis, extracellular matrix formation, glycolysis, water-salt balance, antioxidant system, and the neutrophils involving defense response. It should be noted that levels of histidine triad 1 nucleotide-binding protein 1 (HINT1) and myosin light chain 4 (MYL4) that play an important role in cardiac adaptation to a volume overload were found increased on DI day-3. Two days after DI, proteins did not all had returned to baseline levels. Changes in the level of proteins involved in lipid metabolism were observed only after DI completion.

Posttranslational modifications in fibrinogen resulting from induced oxidation or oxidative stress in the organism can have deleterious influence on optimal functioning of fibrinogen, causing a disturbance in assembly and properties of fibrin. The protective mechanism supporting the ability of fibrinogen to function in ROS-generating environment remains completely unexplored. The effects of very low and moderately low HOCl/- OCl concentrations on fibrinogen oxidative modifications, the fibrin network structure as well as the kinetics of both fibrinogen-to-fibrin conversion and fibrin hydrolysis have been explored in the current study. As opposed to 25 & Mcy;m, HOCl/- OCl, 10 mu M HOCl/- OCl did not affect the functional activity of fibrinogen. It is shown for the first time that a number of Met residues, A alpha Met476, A alpha Met517, A alpha Met584, B beta Met367, gamma Met264, and gamma Met94, identified in 10 mu M HOCl/-OCl fibrinogen by the HPLC-MS/MS method, operate as ROS scavengers, performing an important antioxidant function. In turn, this indicates that the fibrinogen structure is adapted to the detrimental action of ROS. The results obtained in our study provide evidence for a protective mechanism responsible for maintaining the structure and functioning of fibrinogen molecules in the bloodstream under conditions of mild and moderate oxidative stress.

Preeclampsia (PE) is a complex and multifaceted obstetric syndrome characterized by several distinct molecular subtypes. It complicates up to 5% of pregnancies and significantly contributes to maternal and newborn morbidity, thereby diminishing the long-term quality of life for affected women. Due to the widespread dissatisfaction with the effectiveness of existing approaches for assessing PE risk, there is a pressing need for ongoing research to identify newer, more accurate predictors. This study aimed to investigate early changes in the maternal serum proteome and associated signaling pathways. The levels of 125 maternal serum proteins at 11-13 weeks of gestation were quantified using liquid chromatography-multiple reaction monitoring mass spectrometry (LC-MRM MS) with the BAK-125 kit. Ten serum proteins emerged as potential early markers for PE: Apolipoprotein M (APOM), Complement C1q subcomponent subunit B (C1QB), Lysozyme (LYZ), Prothrombin (F2), Albumin (ALB), Zinc-alpha-2-glycoprotein (AZGP1), Tenascin-X (TNXB), Alpha-1-antitrypsin (SERPINA1), Attractin (ATRN), and Apolipoprotein A-IV (APOA4). Notably, nine of these proteins have previously been associated with PE in prior research, underscoring the consistency and reliability of our findings. These proteins play key roles in critical molecular processes, including complement and coagulation cascades, platelet activation, and insulin-like growth factor pathways. To improve the early prediction of PE, a highly effective Support Vector Machine (SVM) model was developed, analyzing 19 maternal serum proteins from the first trimester. This model achieved an area under the curve (AUC) of 0.91, with 87% sensitivity and 95% specificity, and a hazard ratio (HR) of 13.5 (4.6-40.8) with p < 0.001. These findings demonstrate that serum protein-based SVM models possess significantly higher predictive power compared to the routine first-trimester screening test, highlighting their superior utility in the early detection and risk stratification of PE.

Primenenie metodov pryamoj mass-spektrometrii yavlyaetsya odnim iz perspektivnyh podhodov k uluchsheniyu polnoty rezekcii glial'nyh opuholej za schet ispol'zovaniya dopolnitel'nogo sposoba povysheniya tochnosti opredeleniya granic opuholi neposredstvenno v hode nejrohirurgicheskogo vmeshatel'stva. Massivy dannyh, nakaplivaemye v hode aprobacii podobnyh tekhnologij, mogut byt' ispol'zovany i dlya provedeniya fundamental'nyh issledovanij s cel'yu vyyavleniya metabolicheskih izmenenij, soprovozhdayushchih rost opuholi. Cel'yu raboty bylo provesti analiz izmenenij v lipidnom sostave kletochnyh membran diffuznyh i anaplasticheskih astrocitom na osnove dannyh, sobrannyh v hode pryamogo mass-spektrometricheskogo profilirovaniya tkanej, issechennyh v hode planovogo nejrohirurgicheskogo vmeshatel'stva. Lipidnye profili, poluchennye v hode issledovaniya obrazcov opuholevyh tkanej (n = 43) metodom protochnoj mikroekstrakcii v kartridzhe, analizirovali s ispol'zovaniem linejnogo diskriminantnogo analiza so szhatiem, chto pozvolilo vydelit' nabor lipidov, soderzhanie kotoryh izmenyaetsya pri uvelichenii stepeni zlokachestvennosti opuholi. Raznoobrazie lipidov snizhaetsya po mere povysheniya stepeni zlokachestvennosti, tak, soderzhanie 13 fosfolipidov, prinadlezhashchih k 6 razlichnym podklassam, okazyvaetsya snizhennym v anaplasticheskih opuholyah po sravneniyu s diffuznymi. S rostom zlokachestvennosti opuholi umen'shayutsya kak srednij razmer zhirnokislotnyh ostatkov polyarnyh lipidov, tak i stepen' ih nenasyshchennosti. Poluchennye rezul'taty horosho soglasuyutsya s dannymi, poluchennymi ranee v ramkah issledovaniya vysokozlokachestvennyh glial'nyh opuholej, i podtverzhdayut biohimicheskie predstavleniya o pereprogrammirovanii metabolizma v hode malignizacii nejroglii.

BACKGROUND:Schizophrenia is a psychiatric disorder known to affect brain structure and functionality. Structural changes in the brain at the level of gross anatomical structures have been fairly well studied, while microstructural changes, especially those associated with changes in the molecular composition of the brain, are still being investigated. Of special interest are lipids and metabolites, for which some previous studies have shown association with schizophrenia. AIM:To utilize a spatially resolved analysis of the brain lipidome composition to investigate the degree and nature of schizophrenia-associated lipidome alterations in the gray and white matter structures of two neocortical regions - the dorsolateral prefrontal cortex (Brodmann area 9, BA9) and the posterior part of the superior temporal gyrus (Brodmann area 22, posterior part, BA22p), as well compare the distribution of the changes between the two regions and tissue types. METHODS:We employed Matrix-Assisted Laser Desorption/Ionization Mass Spectrometric Imaging (MALDI-MSI), supplemented by a statistical analysis, to examine the lipid composition of brain sections. A total of 24 neocortical sections from schizophrenia patients (n=2) and a healthy control group (n=2), representing the two aforementioned neocortical areas, were studied, yielding data for 131 lipid compounds measured across more than a million MALDI-MSI pixels. RESULTS:Our findings revealed an uneven distribution of schizophrenia-related lipid alterations across the two neocortical regions. The BA22p showed double the differences in its subcortical white matter structures compared to BA9, while less bias was detected in the gray matter layers. While the schizophrenia-associated lipid differences generally showed good agreement between brain regions at the lipid class level for both gray and white matter, there were consistently more discrepancies for white matter structures. CONCLUSION:Our study found a consistent yet differential association of schizophrenia with the brain lipidome composition of distinct neocortical areas, particularly subcortical white matter. These findings highlight the need for broader brain coverage in future schizophrenia research and underscore the potential of spatially resolved molecular analysis methods in identifying structure-specific effects.

Lipids are the most abundant but poorly explored components of the human brain. Here, we present a lipidome map of the human brain comprising 75 regions, including 52 neocortical ones. The lipidome composition varies greatly among the brain regions, affecting 93% of the 419 analyzed lipids. These differences reflect the brain's structural characteristics, such as myelin content (345 lipids) and cell type composition (353 lipids), but also functional traits: functional connectivity (76 lipids) and information processing hierarchy (60 lipids). Combining lipid composition and mRNA expression data further enhances functional connectivity association. Biochemically, lipids linked with structural and functional brain features display distinct lipid class distribution, unsaturation extent, and prevalence of omega-3 and omega-6 fatty acid residues. We verified our conclusions by parallel analysis of three adult macaque brains, targeted analysis of 216 lipids, mass spectrometry imaging, and lipidome assessment of sorted murine neurons. While our brain is primarily composed of lipids, their functions have largely remained unexplored. Here, authors show that specific lipids can be linked to the structural organization and functional hierarchy of the human and macaque brain.

This article studies the structural-functional damage of fibrinogen (FG) treated with hypochlorous acid (HOCl) in the concentration range (10–100 µM). Using tandem mass spectronomy (the MS/MS method), 15 modified amino acid residues with a dose-dependent susceptibility to the oxidizing agent are detected. Using turbidity measurements and confocal laser scanning microscopy (CLSM), it is shown that FG oxidation by 25–100 µM HOCl leads to the formation of a denser fibrin gel, a delayed onset of polymerization, and a decrease in the slope of the polymerization curve, presumably due to the conformational changes in the protein. At lower a HOCl concentration (10 µM), at least six amino acid residues are substantially modified (9–29

The study of urinary peptidome is an important area of research, which concerns the characterization of endogenous peptides, as well as the identification of biomarkers for a wide range of socially significant diseases. First of all, this relates to renal and genitourinary pathologies and/or pathologies associated with proteinuria, such as kidney diseases, bladder, prostate and ovarian cancers, diabetic nephropathy, and pre-eclampsia. Unlike proteins, peptides do not require proteolytic hydrolysis, can be analyzed in their native form and can provide certain information about occurring (patho)physiological processes. Mass spectrometry (MS)-based approaches are the most unbiased and sensitive instruments with high multiplexing capacity and provided most of the current information about endogenous urine peptides. However, despite the large number of urine peptidomic studies, there are certain issues related to the insufficient comparability of their results due to the lack of consistent approaches to their interpretation. Also the development of a custom project-specific protein library for endogenous peptides search and identification is another important point that should be noted in the context of high-throughput peptidomic analysis. Here we propose the custom-specific urinary protein database and the grouping of endogenous urinary peptides with overlapping sequences as useful tools, which can facilitate the acquisition and analysis of LC-MS peptidomic data, as well as the comparison of results of different studies, which should facilitate their more efficient further application.

Accumulation of neurotoxic aggregates of beta-amyloid peptides (Aβ) is a hallmark of Alzheimer's disease (AD) progression. Post-translational modifications (PTMs) increase Aβ aggregation and cytotoxicity, and the content of specific Aβ proteoforms is elevated in senile plaques of AD patients. The pathophysiological mechanisms of aggregate formation and the role of Aβ proteoforms need thorough study both to understand the role played by specific processes in the initiation of neuronal degradation and to find effective preventive means of therapeutic action. The present work investigates the dynamics of accumulation of phosphorylated serine-8 proteoform Aβ (pSer8-Aβ) using the 5xFAD mouse amyloid model. Aβ samples from human cerebrospinal fluid (CSF) and brain were also investigated. Western blot studies using 1E4E11 and 4G8 antibodies showed that accumulation of pSer8-Aβ in mouse brain starts as early as at the age of 3 months and reaches a maximum by the age of 14-17 months, which is generally similar to the dynamics of accumulation of the total pool of Aβ peptides. The pSer8-Aβ level in human CSF in AD patients can reach ~ 1-10% of the total amount of Aβ. Mass spectrometric analysis showed that Aβ phosphorylation by the Ser8, Tyr10, and Ser26 residues in brain tissues, as well as phosphorylation of the APP by Thr719 residue, is possible. These findings support the assumption that pSer8-Aβ proteoforms are involved in amyloidosis in AD. KEYWORDS Beta-amyloid, mass spectrometry, Alzheimer's disease, phosphorylation.

Objectives The COVID-19 pandemic has exposed a number of key challenges that need to be urgently addressed. Mass spectrometric studies of blood plasma proteomics provide a deep understanding of the relationship between the severe course of infection and activation of specific pathophysiological pathways. Analysis of plasma proteins in whole blood may also be relevant for the pandemic as it requires minimal sample preparation.Methods The frozen whole blood samples were used to analyze 203 plasma proteins using multiple reaction monitoring (MRM) mass spectrometry and stable isotope-labeled peptide standards (SIS). A total of 131 samples (FRCC, Russia) from patients with mild (n=41), moderate (n=39) and severe (n=19) COVID-19 infection and healthy controls (n=32) were analyzed.Results Levels of 94 proteins were quantified and compared. Significant differences between all of the groups were revealed for 44 proteins. Changes in the levels of 61 reproducible COVID-19 markers (SERPINA3, SERPING1, ORM1, HRG, LBP, APOA1, AHSG, AFM, ITIH2, etc.) were consistent with studies performed with serum/plasma samples. The best-performing classifier built with 10 proteins achieved the best combination of ROC-AUC (0.97-0.98) and accuracy (0.90-0.93) metrics and distinguished patients from controls, as well as patients by severity.Conclusions Here, for the first time, frozen whole blood samples were used for proteomic analysis and assessment of the status of patients with COVID-19. The results obtained with frozen whole blood samples are consistent with those from plasma and serum.