Synthesis and Characterization of Immunomagnetic Nanoparticles Modified by Monoclonal Antibodies Against Human Breast Mammaglobin
FAN Cai-xia1,2, XU Xin3*, CHEN Zhi-xi2, XIAO Wang-chuan4, JI Xiao-feng5
1. Shantou University Medical College,Shantou 515041, China; 2. Longhua District People��s Hospital, Shenzhen 518109, China; 3. Affiliated Yuebei People��s Hospital, Shantou University Medical College,Shaoguan 512024, China; 4. College of Resource and Chemical Engineering, Sanming University, Sanming 365004, China; 5. Department of General Surgery, Sanming First Hospital,Sanming 365004, China
Abstract��OBJECTIVE To synthesize immunomagnetic nanoparticles with uniform particle size, strong superamagnetism as well as strong immune activity which can be specifically and sensitively combined with circulating tumor cells in peripheral blood of patients with breast cancer.METHODS Superparamagnetic oxide iron nanoparticles containing active carboxyl groups (SMNP-COOH) were synthesized by polyol methods, thermogravimetric analysis was used to determine the amount of carboxyl groups on the surface of SMNP-COOH, while the content of iron was determined by o-phenanthroline. Mediated by 1-ethyl-3,3-dimethylaminopropyl carbodiimide(EDC) and N-hydroxysuccinimide (NHS), immunomagnetic nanoparticles(IMNP) against human breast carcinoma cell line were constructed by binding the monoclonal antibodies against hMAM with SMNP-COOH. X-Ray diffraction was used to confirm their synthesis,meanwhile,transmission electron microscope (TEM), dynamic light scattering (DLS), and vibrating sample magnetometry (VSM) were applied to characterize their physicochemical properties. The conjugation amount of the antibodies and the activity of IMNPs were evaluated by enzyme linked immunosorbent assay (ELISA).RESULTS X-Ray diffraction showed that the chracteristic peaks of the crystalline powder of SMNP-COOH and IMNP agreed with the Fe3O4 standard. The concentration of iron in SMMP-COOH and IMNP were 0.205 and 0.164 mol��L-1, respectively.TEM showed that both synthesized SMNP-COOH and IMNP were almost spherical or ellipsoidal. The sizes of SMNP-COOH and IMNP were (13.7��3.6) and (15.4��4.5) nm, respectively. Dynamic light scattering(DLS) demonstrated the intensity particle size and polydispersity index (PDI) of SMNP-COOH and IMNP were 23.4 nm and 0.303, and 71.2 nm and 0.175,respectively. VSM results showed that both SMNP-COOH and IMNP had strong superamagnetism, and the saturation magnetization of SMNP-COOH and IMNP were 71.37 and 67.68 emu��g-1Fe, respectively, which confirmed antibody binding may reduce the magnetism of SMNP-COOH. The ELISA results showed the conjugation amount of antibody was about 93 ��g on 1 mg SMNP-COOH by covalent bond. The obtained immunomagnetic nanoparticles (IMNP) which were bound with the hMAM monoclonal antibodies could specifically and sensitively combine with breast cancer cell line MDA-MB-415.CONCLUSION IMNP with strong superparamagnetic property,excellent stability and perfect antibody activity were successfully synthesized, which demonstrate the potential to magnetically separate circulating tumor cells in peripheral blood from patients with breast cancer, thus providing a favorable weapon to accurately detect CTCs in breast tumor patients.
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FAN Cai-xia, XU Xin, CHEN Zhi-xi, XIAO Wang-chuan, JI Xiao-feng. Synthesis and Characterization of Immunomagnetic Nanoparticles Modified by Monoclonal Antibodies Against Human Breast Mammaglobin. Chinese Pharmaceutical Journal, 2018, 53(23): 2034-2040.
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