ABSTRACT
Immunogenic antigen (spinosin-BSA) and coating antigen (spinosin-OVA) of spinosin were synthesized by sodium periodate oxidation method. UV scanning analysis method showed that these two spinosins were successfully conjugated with carrier protein and the coupling ratio was 17 and 13.7, respectively. Meanwhile, when immunized by spinosin-BSA，the mice can produce anti-spinosin antibodies with the high titer (1:32 000),specificity (IC₅₀ 211.6 μg·L⁻¹) and low cross-reaction rate measured by ELISA tests. The artificial antigen of spinosin was successfully synthesized, which can be applied for preparation of monoclonal antibodies and establishment of appropriate immune method.
ABSTRACT
<p><b>OBJECTIVE</b>To establish a stable cell line overexpression heme oxygenase-1 (HO-1) mediated by a modified lentivirus system and identify its function.</p><p><b>METHODS</b>The HO-1 gene was amplified by polymerase chain reaction and cloned into the modified pLentiLox3.7 expression vectors. The recombinant plasmids were transfected into HEK293T cells and the HO-1 was detected by Western blot. The recombinant plasmids were transfected into HEK293T cells to produce the viruses, with the helping plasmids including plp1, plp2, and VSVG. HEK293T cells were infected by the viruses and the cells that can express HO-1 were identified by Western blot. The reactive oxygen species were detected in the HO-1-overexpression HEK293T cells and the normal cells after the adding of hydrogen peroxide. The same experiment was performed with human umbilical vein endothelial cells.</p><p><b>RESULTS</b>The stable cell line that can overexpress HO-1 was established, which was verified by Western blot. The reactive oxygen species in the HO-1-overexpression HEK293T cells and human umbilical vein endothelial cells decreased obviously after exposure to hydrogen peroxide.</p><p><b>CONCLUSIONS</b>The lentivirus-carrying HO-1 was successfully packaged and the stable cell line overexpression HO-1 was established. HO-1 can play a protective role in the course of oxidative damage.</p>