Additionally, the most likely reason for a possible false-negative test result is signal suppression by nonspecific immunoglobulins. == Number 3. the assay results. The proposed approach was applied for the serodiagnosis using a recombinant RBD protein of SARS-CoV-2. As a result, an increase in the intensity of test zone coloration by Lanabecestat more than two orders of magnitude was shown, which enabled the significant reduction of false-negative results. The diagnostic level of sensitivity of the LFIA was 62.5% for the common format and 100% for the enhanced format. Moreover, the diagnostic specificity of both variants was 100%. Keywords:immunochromatography, test strips, RBD protein, COVID-19, coronavirus == 1. Intro == Detection of pathogen-specific antibodies in the blood (serodiagnosis) plays a key part in the analysis of many infectious diseases. The advantages of this method are the choice of a specific biomatrix for screening (in contrast with the dedication of viruses or bacteria in different organs and cells), and the similarity of the assay protocols for numerous infections [1]. In medical diagnostics, microplate-based enzyme-linked immunosorbent assays (ELISAs), agglutination Tshr checks, and additional immunoassay types are successfully used for this purpose [2,3,4]. The Lanabecestat current tendency in the development of diagnostic tools is definitely simplification and acceleration of the screening process. The lateral circulation immunoassay (LFIA) is one of the most relevant immunoassay types, with the availability of industrial facilities for the wide developing of diagnostic packages. Multimembrane composites (test pieces) with initial applied immune reactants and coloured nanoparticles as recognized labels provide maximum reducing manipulations of an operator. Contact of a test strip with the tested sample initiates the lateral circulation of the immune reactants along the membranes and the assay results can be visualized within 1015 min from the coloration of the test strip zones [5]. Successful development of the LFIA-based serodiagnosis of various diseases is definitely presented in several publications [6,7,8,9,10]. As an additional application, the assessment of the vaccine performance and the organisms protection against infections can be described [11,12]. The recent COVID-19 pandemic also caused the necessity of serodiagnostic LFIAs, which were successfully developed and commercialized by different organizations and companies [13,14]. However, the development and practical application of a serodiagnostic LFIA are accomplished by significant limitations. The percentage of the illness instances revealed from the LFIA serodiagnosis is definitely often inferior to the instances recognized by instrumental immunoassays, such as the ELISA. The main factor causing this obstacle is the need to detect specific antibodies with low concentrations in the presence of great excess of total immunoglobulins [15]. Serodiagnosis is typically implemented using immunoglobulin-binding proteins, such as anti-species antibodies or bacterial protecting compounds (staphylococcal protein A, streptococcal protein G, etc.) [16]. In the common LFIA file format, they may be immobilized on colored nanoparticles, applied on a test strip, and form complexes with all immunoglobulins, including those specific to the prospective pathogen Lanabecestat during lateral circulation. The producing complexes interact with the antigen immobilized in the test zone (TZ) of the strip, whereas unbound compounds move across this zone. Therefore, the coloration of the TZ shows the presence of specific antibodies in the tested sample [8,17,18,19]. However, nonspecific immunoglobulins block binding sites at the surface of nanoparticles and only a minor portion of specific antibodies can form labeled complexes in the TZ. This differs from your ELISA-based serodiagnosis where non-bound antibodies can be eliminated (washed) prior to the inclusion Lanabecestat of labels into immune complexes. This limited binding prospects to a significant percentage of false-negative results, which are often indicated during the validation of these assays [20,21]. The possibility to change the order of the immune complexes formation was regarded as in alternate serodiagnostic LFIAs [20,22,23,24]. As a result, some limitations were mentioned in these methods. For example, the Lanabecestat inverted LFIA file format, where the antigen is definitely immobilized on label particles and the immunoglobulin-binding protein is definitely adsorbed within the operating membrane, allows for the binding of more immunoglobulins than the traditional LFIA file format. Nevertheless, in this case, the interference of non-specific immunoglobulins is not eliminated [25,26,27]. One of the approaches to get rid of their influence is the double antigen sandwich LFIA format based on the antibody polyvalence [23,28,29,30]. However, due to the part formation of polyvalent complexes, the effectiveness of this format strongly depends on the percentage of the reagents [15]. Moreover, the.