ABSTRACT
BACKGROUND: The unique expression pattern of prostate stem cell antigen (PSCA) in a number of prevalent neoplasms has made the antigen a great target for cancer researches, and many clinical methods have been developed based on the application of this tumor marker. Hence, optimal PSCA laboratory production can be considered a hallmark for many researchers. OBJECTIVE: An analytical study was designed to improve the quality and quantity of PSCA production. MATERIALS AND METHODS: The effects of different compositions of lysis buffers and some ultrasound durations were assessed by calculation of the protein recovery followed by PSCA specific blotting experiments. Then, based on the results of the web-based characterization, interference removal, followed by re-solubilization of the protein in various buffers, was designed, applied, and assessed. RESULTS: Since the selection of an appropriate methodology depends merely on the research purposes, we tried to discuss the pros and cons of the investigated methods according to the hydrophobic nature of PSCA as well as its dramatic tendency to aggregate in the form of inclusion bodies in the expression hosts. CONCLUSIONS: We introduced a newly designed method to fit the delicate immunological surveys and overcome some limiting factors in PSCA production.
ABSTRACT
Background: The main property of a successful conjugation of antibodies to nanoparticles is keeping the potency of antibody for binding the antigen, and an oriented conjugation can do that. Under such ground, this study was carried out to explore the efficiency of two conjugation methods in binding iron nanoparticles to an antibody produced against PSCA (prostate stem cell antigen) using in vitro labeling of PC3 cells. Methods: In this experimental study, we conjugated dextran-superparamagnetic iron oxide nanoparticles (dexSPIONs) to anti-PSCA antibody by two different methods, including targeting carbohydrate moieties in FC domain and the free amine group of amino acid side chains. Ultimately, Iron staining was done by anti-PSCA antibody-dexSPIONs in PC3 cells to detect antibody binding to the cells. Results: A strong blue dye was induced by iron staining in conjugated dexSPIONs on the membrane of PC3 cells by the former method than the second one. Moreover, cells treated with 20 nm diameters of dexSPIONs showed higher resolution of blue color than those treated with 100 nm nanoparticles. Conclusion: This oriented conjugation method promoted the efficiency of targeting tumor antigens, and the presence of iron particles might enhance MRI image intensity in vivo by targeting PSCA-overexpressing cells in future studies.
