Nowadays, the presence of pathogen-specific antibodies in the blood is widely controlled by a serodiagnostic technique based on the lateral flow immunoassay (LFIA). However, its common one-stage format with an antigen immobilized in the binding zone of a test strip and a nanodis-persed label conjugated with immunoglobulin-binding proteins is associated with risks of very low analytical signals. It is caused by the presence of non-specific immunoglobulins in very large excess to the target antibodies in the tested samples thus decreasing their binding with the detected labels. In this study, the first stage of the immunochromatographic serodiagnosis was carried out in its traditional format using a conjugate of gold nanoparticles with staphylococcal immunoglobulin-binding protein A and an antigen immobilized on a working membrane. At the second stage, a labeled immunoglobulin-binding protein was added, which enhanced the coloration of the bound immune complexes. The use of two separated steps, binding of specific antibodies, and further coloration of the formed complexes allowed a significant reducing the influence of non-specific immunoglobulins on the assay results. The proposed approach was ap-plied for the serodiagnosis using a recombinant RBD protein of SARS-CoV-2. As a result, an in-crease in the intensity of test zone coloration by more than two orders of magnitude was demonstrated, which enabled to significantly reduce false-negative results. When testing a panel of 16 positive and 8 negative serum samples, the diagnostic sensitivity of the LFIA was 62.5% for the common format and 100% for the enhanced format; the diagnostic specificity of both variants was 100%.

(1) Background: There is a risk of Aflatoxin B1(AFB1) pollution in yellow rice wine[1],threatening human life and health, the risk is a major focus of government attention and social concern.The current detection technology cannot meet the needs of large-scale testing due to the disadvantages of high false positive, cumbersome operating procedures or high cost.Therefore, it is urgent to study and establish a high-sensitivity and high-fast detection method for AFB1 to provide technical support for yellow rice wine quality supervision and risk assessment.; (2) Methods: In this study, three-factor and three-level orthogonal experiments were carried out for oscillation time, reaction temperature and tomography time to determine the best reaction conditions for pretreatment.AFB1 detection was carried out by aflatoxin time-resolved fluorescence immunochromatography technology，Through the ratio of the T signal value of the detection line to the C signal value of the quality control line and the natural logarithmic value of the standard solution concentration[2], a rapid detection technology of time-resolution fluorescence immunochromatography is established.The technology was methodically assessed, the actual yellow rice wine samples were tested, and the results were compared with the HPLC method; (3) Results: The results show that the optimal reaction conditions for pretreatment are: the sample oscillation time is 20min, the sample reaction temperature is 37°C, and the sample chromatography time is 6min. In the actual sample detection of yellow rice wine, The time-resolved fluorescence immunochromatography detection technology established in this study detects AFB1. The detection limit is 0.3 μg·kg-1,The linear range is 0.8-12.0μg·kg-1,The linear equation of the corresponding standard curve y=-0.1913x+0.5206 (R2=0.9948),The standard deviation is 0.029.The results of the in-batch accuracy and precision of this method show that the addition and recovery rate is 77.9% - 105.7%,The coefficient of variation is in 4.5% - 10.3%; Inter-batch accuracy and precision test results show that the addition recovery rate is75.9% - 85.8%,the coefficient of variation is in 8.1% - 13.9%,explain that the detection technology has good accuracy and precision.Compared with HPLC, the relative error is less than 10%,It shows that aflatoxin time-resolution fluorescence immunochromatography technology is in good agreement with the test results of national standard methods; (4) Conclusions:he rapid detection technology of time-resolution fluorescence immunochromatography of AFB1 in yellow rice wine is easy to operate with fast detection speed, high sensitivity, good repeatability and stability, and has broad application prospects.

The presented study is focused on the impact of binding zones locations at immunochromatographic test strips into analytical parameters of multiplex lateral flow assay. Due to non-equilibrium conditions for such assays the duration of immune reactions influences significantly on analytical parameters, and the integration of several analytes into one multiplex strip may cause essential decrease of sensitivity. To choose the best location of binding zones, we have tested reactants for immunochromatographic assays of lincomycin, chloramphenicol, and tetracycline. The influence of the distance to the binding zones on the intensity of coloration and limit of detection (LOD) was rather different. Basing on the obtained data, the best order of binding zones was chosen. In comparison with non-optimal location the LODs were 5-10 fold improved. The final assay provides LODs 0.4, 0.4 and 1.0 ng/mL for lincomycin, chloramphenicol, and tetracycline, respectively. The proposed approach can be applied for multiassays of other analytes.

Determination of the presence in the blood of antibodies specific to the causative agent of a particular disease (serodiagnosis) is an effective approach in medical analytical chemistry. Serodiagnostics performed in the lateral flow immunoassay format (immunochromatography) meet the modern requirements for point-of-care testing and are supported by existing technologies of large-scale diagnostic tests production—thus raising increased attention in a tense epidemiological situation. For traditional lateral flow serodiagnostics formats, a large number of nonspecific immunoglobulins in the sample significantly reduces the degree of detectable binding. To overcome these limitations, an assay based on the formation of immobilized antigen— specific antibody—labeled antigen complexes detection was proposed. However, the requirements for its implementation, providing maximum sensitivity, have not been established. This article describes the mathematical model for the above assay. The influence of the ratio of reagent concentrations on the analysis results is considered. It is noted that the formation of specific antibody complexes with several labeled antigens is the main limiting factor in reducing the detection limit, and methods are proposed to minimize this factor. Recommendations for the choice of the assay conditions, following from the analysis of the model, are confirmed experimentally.