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The lack of specific clinical symptoms for patients in the early stage of rheumatoid arthritis (RA) has created strong interest in the laboratory diagnosis of RA. The main laboratory markers of RA, rheumatoid factor (RF) and anti-citrullinated protein antibodies (ACPAs), can be found in patients with other pathologies and in healthy donors. Even today, there is no single laboratory test that can diagnosis RA with high sensitivity and specificity. To improve the diagnosis and treatment of RA, alternative biomarkers, including 14-3-3η protein, connective tissue growth factor (CTGF), antibodies against PAD4, antibodies against BRAF, and anti-acetylated and anti-carbamylated protein antibodies have been studied extensively. The use of a multiple biomarker approach, the simultaneous measurement of a set of biomarkers, is an alternative strategy for the diagnosis of RA and for predicting the therapeutic effect of biological disease-modifying antirheumatic drugs (DMARDs). However, despite the large number of studies, only a few biomarker combinations have been validated and can be applied in clinical practice. In this article, results of studies focused on the multiple biomarker approach (both multiplex and combined single-analyte assays) to diagnose RA and to predict response to biological drug therapy are reviewed. Additionally, general factors limiting the use of multiplex analysis in RA diagnostics and therapy are discussed.

Cytokines and acute phase proteins play an important role in the development of the immune response during inflammatory reactions. Depending on the type of disease, the development of inflammation is accompanied by changes in concentrations (both decrease and increase) of not one, but many inflammatory biomarkers. Here, a quantitative microarray-based method for multiplex immunoassay of eight biomarkers of human inflammation, namely acute phase proteins (C-reactive protein, serum amyloid protein A) and cytokines (IL-6, IL-8, IL-17, IL-18, IP10/CXCL10, TNFα) was developed and the possibility of its use for the detection of inflammatory biomarkers in a culture medium has been demonstrated. The developed method can be used to evaluate changes of the inflammatory biomarker profile induced by different agents or to determine the concentrations of biomarkers after activation of cells while studying different diseases with the help of in vitro models.

Context Autoimmune polyglandular syndrome (APS) is a cluster of endocrine disorders arising from immune dysregulation, often combined with damage to nonendocrine organs. There are 2 types of APS: type 1 and type 2 (APS-1 and APS-2, respectively). In clinical practice, an atypical course of APS is often observed. Objective This work aims to find a novel genetic predictor of APS. Methods We performed exome sequencing in 2 patients with an atypical clinical APS picture and members of their families. Patient A presented with a manifestation of APS-2 in early childhood and patient B with a late manifestation of the main components of APS-1. Results In patient B, we identified inherited compound mutations as a novel combination of the c.769C > T and c.821delG alleles of AIRE and genetic variation in the CIITA gene. No homozygous or compound mutations in AIRE were found in patient A, but we did reveal mutations in genes encoding regulatory proteins of innate and acquired immunity in this patient. Conclusion Our data revealed novel combination of mutations in the AIRE gene in atypical APS and imply that mutations in immune-related genes may modify the clinical manifestation of APS in AIRE-mutation carriers and contribute to the development of autoimmune pathology in non-AIRE carriers with atypical APS.

The diagnosis of autoimmune polyglandular syndrome (APS) types 1/2 is difficult due to their rarity and nonspecific clinical manifestations. APS-1 development can be identified with assays for autoantibodies against cytokines, and APS-2 development with organ-specific antibodies. In this study, a microarray-based multiplex assay was proposed for simultaneous detection of both organ-specific (anti-21-OH, anti-GAD-65, anti-IA2, anti-ICA, anti-TG, and anti-TPO) and APS-1-specific (anti-IFN-ω, anti-IFN-α-2a, and anti-IL-22) autoantibodies. Herein, 206 serum samples from adult patients with APS-1, APS-2, isolated autoimmune endocrine pathologies or non-autoimmune endocrine pathologies and from healthy donors were analyzed. The prevalence of autoantibodies differed among the groups of healthy donors and patients with non-, mono- and multi-endocrine diseases. APS-1 patients were characterized by the presence of at least two specific autoantibodies (specificity 99.5%, sensitivity 100%). Furthermore, in 16 of the 18 patients, the APS-1 assay revealed triple positivity for autoantibodies against IFN-ω, IFN-α-2a and IL-22 (specificity 100%, sensitivity 88.9%). No anti-cytokine autoantibodies were found in the group of patients with non-APS-1 polyendocrine autoimmunity. The accuracy of the microarray-based assay compared to ELISA for organ-specific autoantibodies was 88.8–97.6%. This multiplex assay can be part of the strategy for diagnosing and predicting the development of APS.

Colorectal cancer (CRC) is one of the most common primary malignancies. Early stages of the disease are asymptomatic in the majority of cases, leading to late detection and high mortality. Available noninvasive diagnostic techniques are limited in sensitivity and specificity, and designing new ones is still a pressing problem. Exosomes are membrane-derived microvesicles secreted into human biological fluids and provide a novel way to assess the course of an oncology disease. The review describes the repertoire of exosomal surface biomarkers found in the blood of CRC patients and the prospects of employing multiplexed tests for exosomal markers in early noninvasive diagnosis of cancer.

Community-acquired pneumonia (CAP) is a dangerous disease caused by a spectrum of bacterial and viral pathogens. The choice of specific therapy and the need for hospitalization or transfer to the intensive care unit are determined by the causative agent and disease severity. The microbiological analysis of sputum largely depends on the quality of the material obtained. The prediction of severity and the duration of therapy are determined individually, and existing prognostic scales are used generally. This review examines the possibilities of using specific serological biomarkers to detect the bacterial or viral aetiology of CAP and to assess disease severity. Particular emphasis is placed on the use of biomarker signatures and the discovery of biomarker candidates for a single multiplex analysis.

We propose an approach that allows simultaneous determination of the levels of M and G isotypes of antibodies to the panel of glycans using microarrays. The level of IgG antibodies to 3’-O-su-Lea glycan detects patients with colorectal cancer with a sensitivity of 69.77% and specificity of 62.75%. The percentage of correctly classified colorectal cancer patients with the use of a combination of two markers IgM antibodies to glycans 3’-sialyl-TF and 3’-O-su-Lea is as high as 74.23%. The levels of IgM antibodies to 3’-O-su-Lea glycan differ significantly in patients with and without regional metastases. The levels of some of antiglycan IgM or IgG antibodies differed significantly in patients with tumors of different location and differentiation.

A prototype of test-system for simultaneous quantitative assay of nine tumor markers in blood serum was developed. The main constituent of the test-system is OM-9 biochip containing immobilized antibodies against nine oncomarkers: α-fetoprotein (AFP), carcinoembryonic antigen (CEA), human chorionic gonadotropin (HCG), cancer antigen 15-3 (CA 15-3), cancer antigen 125 (CA 125), cancer antigen 19-9 (CA 19-9), prostate-specific antigen, total (PSAtot) and free (PSAfree) forms, neuron-specific enolase (NSE). The biochip-based assay procedure for carrying out simultaneous quantitative determination of nine tumor markers in patient's blood serum: two-steps sandwich-immunoassay, was proposed. The main analytical characteristics of the method were obtained. The results permit to consider the prototype of the test-system as a promising instrument for clinical application. The test-system prototype was tested using blood serum samples of oncological patients (252 samples) and healthy donors (185 samples). Increased concentrations of one or more tumor markers above the normal level were found in 76.6% cases of oncological patients and only in 6% cases of healthy donors. For colorectal cancer patients group, application of modern statistical methods of data-processing in medical researchers, i.e. ROC-analysis and logistic regression, indicted that the simultaneous assay of nine tumor markers on biochips showed much more diagnostic significance (area under the ROC-curve (AUC) reached 0.84) than traditional assay of 2 tumor markers, CEA and CA 19-9 (AUC = 0.59). The developed biochip-based test-system can be recommended both for the estimation of people's health, e.g., for standard medical examination, and for tracking of tumoral process in postsurgical period or after specific tumor treatment.