Abstract <p>The effect of gold nanoparticle functionalization based on the high-affinity binding of biotinylated antibodies to streptavidin on the surface of gold nanoparticles, which ensures oriented arrangement of the antibodies, as well as the effect of the size of the nanolabel on the sensitivity of immunochromatographic assay, is studied. The object of study is the anti-Müllerian hormone, a biomarker of the state of the female reproductive system. Two methods of antibody immobilization are considered: high-affinity streptavidin–biotin binding and physical adsorption. Oriented immobilization demonstrates the absence of nonspecific interactions and an increase in assay sensitivity. An increase in the size of nanoparticles in the immunoconjugate leads to a decrease in the limit of detection. The identified patterns make it possible to develop a test system with a hormone detection limit of 0.1 ng/mL and a working range of 1.0–42.1 ng/mL. The possibility of using the test system in fivefold diluted blood serum is demonstrated.</p>