Comparison of different antibodies for enrichment myeloid-derived suppressor cells from bone marrow of orthotopic liver tumour mice by using flow cytometry
Received:March 28, 2013  Revised:May 21, 2013
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KeyWord:myeloid-derived suppressor cells,hepatocellular carcinoma,fluorescence-activated cell sorting
赵文秀 厦门大学附属中山医院
张正奇 厦门大学附属中山医院
许雅苹 厦门大学附属中山医院
尹震宇 厦门大学附属中山医院
吴端 厦门大学附属中山医院
王效民 厦门大学附属中山医院
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      【Abstract】 Objective Accumulating evidences have demonstrated that myeloid-derived suppressor cells (MDSCs) play a pivotal role in tumor associated immune suppression. The main purpose of this study was to compare different antibodies to isolate MDSCs form bone marrow of orthotopic liver tumour mice by flow cytometry. Methods The in situ hepatic tumour model was created by the direct intrahepatic injection of mouse hepatoma cells lines H22 (1×106 cells originating from the BALB/c mice). Then the mice were sacrificed in 10 days for anatomy. The monouclear cells were isolated from bone marrow and stained with CD11b,Gr-1 or double stained with CD11b and Gr-1 and then sorted by flow cytometry. The expression of arginase-1(Arg-1) and inducible nitricoxide synthase (iNOS)of isolated MDSCs were detected by flow cytometry. Cellular reactive oxygen species(ROS) detection assay kit was used to measure ROS production by MDSCs. Results The purity and viability of isolated MDSCs by different antibodies was all greater than 90%. Enrichment of MDSCs by CD11b antibody is much more easy, but relative low purity. MDSCs sorted by Gr-1 antibody have higher purity, but the cell population is hard to definite. MDSCs isolated by double staining have the highest purity. MDSCs express high levels of Arg-1, iNOS and ROS. Conclusion High purity and viability of MDSCs can be sorted from bone marrow of hepatic tumour bering mice by different labelling methods.Enrichment of MDSCs by CD11b is easy to operate, unified and copy, which can be served as the first choice in sorting MDSCs . The successful sorting of MDSCs was important for the further studing the biological and immunological function of MDSCs.
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