ABSTRACT
BACKGROUND: The imaging technique of CT and MRI has lower diagnostic accuracy in analyzing lymph node location due to the nonspecific diagnostic criteria. However, this situation can be improved with Gadolinium diethylenetriamine pentaacetic acid (Gd-DTPA), especially the use of lymphatic targeting contrast agent. OBJECTIVE: To prepare DTPA-coupled human serum albumin (HSA) labeled with gadoliniumion, and to investigate the enhancement for the lymph node in animal experiment, in further, to explore the potential value of its application. DESIGN, TIME AND SETTING: The randomized grouping, controlled animal experiment was performed at the laboratory of Medical Imaging Center, Nanfang Hospital from December 2008 to April 2009. MATERIALS: GdCI3 was prepared with Gd2O3 reacted with 0.1mol/L HCI for 10 minutes. Then, DTPA cyclic anhydride and HAS solution were mixed together at the ratio of 200:1 or 100:1. The product was divided into 4 equal portions, which was reacted with 5.0, 6.0, 7.0, 8.0 pH values at room temperature for 30 minutes. At last, GdCI3 was added. The complexation reaction was completed within 1 minute. The Gd-DTPA-HAS was dried and prepared for Gd-DTPA-HAS solution with concentration of 0.50 mmol Gd/L.METHODS: Six New Zealand rabbits were undergone the MRI examination, followed by subcutaneous injection of 0.2 mL with Gd 0.50 mmol/L HSA-Gd-DTPA solution. And contrast scanning was performed at minutes 30, 60 and 180 after injection. The signal intensity of popliteal lymph node was measured and the enhancement rate was obtained. MAIN OUTCOME MEASURES: The number of combined Gd-DTPA per HSA at different reaction conditions was measured, and coupling efficiency of HSA-DTPA was calculated. The enhancement rates of lymph node, as well as dynamic changes were observed. RESULTS: ①The number and coupling efficiency of HSA-coupled of gadolinium ions were increased with the pH value of the reaction system at the same DTPA-to-HSA molar ratio. ②At the same pH value of the reaction system, the number of HSA-coupled of gadolinium ions increased with the DTPA-to-HSA molar ratios while the coupling efficiency of HSA-coupled of gadolinium ions decreased.③After subcutaneous injection, HSA-Gd-DTPA entered the lymphatic vessel, which resulted in obvious homogeneous enhancement of the lymphatic vessels and popliteal lymph nodes. CONCLUSION: The number and coupling efficiency of HSA-coupled of gadolinium ions can be increased in pH=8.0, 0.1 mol/L NaHCO_3 reaction system. HSA-Gd-DTPA is an effective contrast agent targeted for lymph node after subcutaneous administration.
ABSTRACT
Objective To evaluate the enhancing effect of MRI with Gd DTPA F(ab)′ 2 in nude mice models with LS 174T tumor, and to investigate the influence of Gd labeling on the immunocompetence of the fragments (ab)′2 of CL 3. Methods (1)Antibodies and tumor models preparation: Monoclonal antibodies (McAb) fragment F(ab)′ 2 against LS 174T colonic adenocarcinoma cell were prepared with hybridoma cell CL 3. Twenty BALB/C nude mice were used as tumor models by subcutaneous injection of LS 174T cells. (2)Gd 3+ Labelling: DTPA anhydride and F(ab)′ 2 fragments were conjugated. Two groups were classified: group A, with C/P=2∶1; group B, with C/P=5∶1. Then, Gd was added into DTPA F(ab)′ 2 solution. Gd DTPA F(ab)′ 2 was formed. Free Gd 3+ and DTPA were removed by dialysis. Purified Gd DTPA F(ab)′ 2 was available. (3) MR scanning techniques: MRI was performed with a GE Signa Horizon LX MR scanner. The nude mice were immobilized and divided into three groups: routine plain scan group, enhanced scan group A(C/P=2∶1)and group B(C/P=5∶1). Results When pH=8.0 ,[F(ab)′ 2]=4.6 mg/ml, and C/P=2∶1 or C/P=5∶1, immuno conjugated rates of group A and B were 74% and 62%, respectively. Immunocompetence of fragment F(ab)′ 2 reduced approximately 12% when C/P increased from 2∶1 to 5∶1.The enhancing effects of two groups of nude mice were both poor, and that of group B(C/P=5∶1) was better than that of group A(C/P=2∶1). Conclusion While C/P increasing, immunocompetence of fragment F(ab)′ 2 reduced, and the enhancing effects of group B was better than group A. The enhancing effect of Gd DTPA F(ab)′ 2 was tumor specific.
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Objective To further improve the amount of gadolinium located on tumor, a gadolinium chelate enhanced magnetic resonance imaging pretargeting with avidin-biotin system technique was adopted and the enhancing characteristics of difference of signal intensity at various scan timing were investigated in our experiment. Methods (1) Anti-CEA antibody CL -3 was biotinylated in a mixture with antibody to NHS-LS-biotin with a molar ratio of 1/30-50. (2) After the reaction of GdCl 3 and DTPA-B, the unconjugated gadolinium was removed by chromatography on G-10 column. (3) Steps for pretargeting tumor: First step, McAb-B was injected intravenously into nude mice on the first day. Second step, avidin (Av) and streptavidin (SA) were injected intraperitoneally 24 hours later. Third step, Gd-DTPA-Bt was injected intravenously 48 hours after the first injection. MRI was performed with plain scans, enhanced scans at 20 minutes, 2 hours, 8 hours, and 24 hours after the third step. Signal intensities of tumor and muscles were measured. The pretargeting effect was compared with those of Gd-DTPA-McAb and Gd-DTPA. Results (1) Each monoclonal antibody conjugated with 11-23 biotin and the immunoactivity of biotinylated antibody with 12 biotin/antibody was 94.9%. (2) The enhancing effect of pretargeting approach was tumor specific. Contrarily that of Gd-DTPA was not. (3) The enhancing rate of signal intensity specificity of pretargeting approach was 43%, while that of McAb-Gd-DTPA was 17.9% only, so the enhancing ratio was 2.4. Conclusion Pretargeting approach using avidin -biotin system improves the amounts of gadolinium locating on tumors and yields a specific enhancing effect. It is a promising modality which promotes the ability of Gd labeled magnetic resonance immunoimaging in the detection of colon cancer and its recurrence.
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Objective To investigate the value of superparamagnetic iron oxide particles(SPIO) in MR imaging for differentiation of metastatic lymph nodes from normal or reactively hyperplastic lymph nodes. Methods For the time course study,10 ?mol Fe of SPIO per extremity was administered to 6 animals, and images were obtained before and 1 h to 48 h after administration. Reactively hyperplastic lymph node model was established in 6 animals by injection of egg yolk emulsion, and tumor bearing lymph node model was established in another 6 rabbits. MR images of the popliteal lymph nodes were obtained before and 12 h after subcutaneous administration of SPIO. Imaging results were analyzed and correlated with pathological findings. Results In unenhanced images, the signal intensity of tumor free and metastatic lymph nodes did not differ significantly. After administration of SPIO, a significant signal intensity loss occurred in the tumor free lymph node, and the maximal loss occurred at 12 h, and the signal intensity reduced by 49%,28%,59%, and 89% on T 1WI, T 2WI, PDWI, and T 2WI, respectively at 48 h( P =0.000). However, the signal intensity of the metastatic lymph nodes remained unchanged. Conclusion MR imaging after subcutaneous administration of SPIO help to differentiate metastatic from normal or reactively hyperplastic lymph nodes.