Résumé
Objective To develop an ultrasmall superparamagnetic iron oxide(USPIO)based MR probe targeting CD40 mutant and investigate its biological and chemical properties and its targeting effect on ovarian cancer cells in vitro.Methods To prepare immunologically competent probe,the monoclonal antibody was conjugated with USPIO particles modified by DMSA based on chemical crosslinking method.The USPIO labeled anti-human CD40 mutant monoclonal antibody 5H6(5H6-USPIO)was the experimental probe,and the USPIO labeled anti-human CD40 monoclonal antibody 5C11(5C11-USPIO)and USPIO served as control agents.The flow cytometry,confocal microscopy and Prussian blue staining were employed to assess the magnetic performance and analyze its bioactivity of the probe.The probe's cell MR imaging in vitro was carried out using ovarian caner cells(HO8910)with high CD40 mutant expression.The analysis of signal data of different groups was conducted by using one-way ANOVA andLSD test.The probe's effect on ovarian caner cells' growth was measured by CCK-8 kit.Results The stable molecular probe carrying nanoparticles and CD40 mutant antibody was built and purified successfully.The probe had similar magnetic property compared with original USPIO.Immunofluorescence and Prussian blue staining confirmed that the molecular probe could recognize CD40 mutant on ovarian cancer cells(HO8910)with high specificity.The probe had no effect on the growth of HO8910 cells.MR cell imaging in vitro showed that the value of T2 and T2 * decreased significantly after the probe binding with HO8910 cells and T2WI became darker than control groups.The T2 and T2* relaxation time of 5H6-USPIO group was(40.05 ± 1.62)ms and(3.08 ± 0.11)ms,respectively.The T2 and T2 * relaxation time of 5H6-USPIO group was shorter than 5C11-USPIO [(85.38 ± 4.74)and(11.82 ± 1.00)ms,respectively]and USPIO[(91.62 ± 3.35)and(13.60 ± 1.92)ms,respectively]groups with statistical significance(F =196.29,60.73,P ＜0.01).The difference of T2 and T2* relaxation time between 5C11-USPIO and USPIO groups showed no statistical significance (P ＞0.05).Conclusions CD40 mutant antibody can conjugate with ultrasmall superparamagnetic iron oxide particles to form MR molecular probe using chemical crosslinking method.The probe has good magnetic characters and high bioactivity.The probe can specifically bind with HO8910 cells.