## 3 Results and Discussion
### ICA on Cattle Sera with QD Amplification  
ICA was implemented according to the common scheme of ICA serodiagnostics [40]. According to this scheme, when a liquid sample migrates along a test strip, specific immunoglobulins interact with the marker-protein G conjugate. Subsequent detection of the formed complex is carried out by binding of the obtained complex with immobilized LPS in the analytical zone. All immunoglobulins in the sample can bind to the marker-protein G conjugate, and the number of specific antibodies capable of forming complexes with both the conjugate of GNPs and LPS in the analytical zone is tens of times lower than the total amount of immunoglobulins. For this reason, the ICA sensitivity is often insufficient for reliable serodiagnostics. In this work, we included in the ICA scheme an additional stage of adding the QD-(protein G) conjugate. Two processes take place in this case: (1) Interaction of the QD-(protein G) conjugate with specific immunoglobulins that did not bind to the GNP-(protein G) conjugate, but bound to LPS in the analytical zone and (2) interaction of the nonspecific immunoglobulins bound to the GNP-(protein G) conjugate in the analytical zone with the QD-(protein G) conjugate. That is, in this case, nonspecific immunoglobulins do not interfere with the assay as they do in the common ICA scheme but, on the contrary, play the role of additional binding sites in the analytical zone. This process results in an increase in signal intensity, which helps obtain more reliable results. The proposed scheme for immunochromatographic serodiagnostics is presented in Figure 3.  
this test system was 95% of that of ELISA. Using the TotalLab software, the images of the test systems were digitized, which allowed the obtaining of quantitative data on the color intensity in the control and analytical zones. As can be seen from the photographs of the test strips after testing four samples with the lowest antibody titers (Figure 4), the GNP-based ICA system showed negative results for two of the four weakly positive sera. A further two sera showed weak staining and were characterized as conditionally positive. All sera that produced staining of the analytical zone of fewer than 0.2 units were considered negative. A signal from 0.2-1 unit was considered conditionally positive. Using the test system with QD, 81 out of 83 sera were identified as positive, and two gave questionable results. That is, the sensitivity of the system that uses QDs as a marker and fluorescence registration of the signal increased to 98% of the sensitivity of ELISA. Finally, for a system with two labels-GNPs and QDs-the sensitivity was not inferior to ELISA, and all 83 samples were identified as positive. To establish the limit of detection for developed test-system 1 \(\upmu\)L of standard serum sample was diluted with 1% PBS containing 1% Triton X-100 from 1:100 to 1:30,000 and then tested using GNP-based and GNP+QD-based immunomodromagnatographic test-systems. As shown in Figure 5, the detection limit (the disappearance of color in the analytical zone) of GNP-based tests accords to dilution of standard serum sample 1:10,000, whereas, for GNP+QD-based tests it is equal to 1:20,000. Figure 4: Results of immunomodromagnatographic serodiagnostics of cattle brucellosis using the GNP-(protein G) conjugate (**A**); using the QD-(protein G) conjugate (**B**); and using the signal amplification with the QD-(protein G) conjugate (**C**). AZ—analytical zone; CZ—control zone. The obtained data confirm that the sensitivity of the assay is increased if a stage of binding with QDs is included. The results presented in Table 1 show that when two stages of signal generation, with the GNP conjugate and the QD conjugate, are used, the signal intensity in the analytical zone increases by 2-3 times compared with the test system based only on the QD conjugate and by more than nine times compared with the system based on the GNP conjugate.