Preparation, Identification And Preliminary Clinical Application Of Polyclonal Antibody Of Atra-related Resistance Gene HA117

Objective: Prepare polyclonal antibody of HA117 in order to explore its distributive characteristics and level of expression in tumor and normal tissues.Methods: By using pAdTrack-HA117 plasmid as the template, PCR amplifies the total cDNA of HA117. The products of PCR were transferred into BL21 (DE3) competent cells to induce the protein expression of HA117, which will be identified by SDS-PAGE. After HA117 protein is purified, it is used to immune New Zealand rabbits to produce polyclonal antibody of HA117. Through indirect ELISA, the titer of the polyclonal antibody of HA117 can be detected. While using HEK293 cell with transducted HA117 gene as the experiment group, HEK293 cell as the comparison group, immunofluorescence and Western blot are used to detect HA117 protein, and to verify the specificity of polyclonal antibody of HA117. Results: Polyclonal antibody of HA117 was got successfully. The anti-HA117 serum titer was 1:729000, showed by indirect ELISA. The HA117 protein in transducted HEK293 can be specifically identified by the polyclone antibody of HA117, showed by immunofluorescence and Western bolt.Conclusion: The acquirement of the polyclonal antibody of HA117, with a high titer and specificity, provided a useful and strong tool for exploring HA117's distributive characteristics and level of expression in tumor and normal tissue, as well as provided a good way for further functional research of HA117. Objective: To study the expression and clinical significance of HA117 proteins in neuroblastoma.Methods: Using 20 cases neuroblastoma as the research object, taking 14 cases peritumoral organization as a comparison, observing them expression by immunohistochemistry method. According to the Shimada pathologic stage method, the neuroblastomas were divided to Favorable Histology Group (FH) and Unfavorable Histology Group (uFH), in order to detect the different expression of HA117. According to whether having lymph node and distant metastasis, dividing into two group to analysis the different expression of HA117 protein. Detecting the different expression and analyzing the clinical significance of MDR1 and HA117 in 20 cases of neuroblastoma .Results:The expression rate of HA117 in neuroblastoma was 70% (14/20), in peritumoral organization was 21% (3/14), the result has statistically significance.The positive rate of FH was 50%, and the positive rate uFH was 100%,higher than FH.Who has lymphatic and distant metastasis were 11 cases, and the positive rate was100%, no transfer cases were 9, and 3 of them had expressed, expression rate was 33%. The result has statistically significance.The expression rate of HA117 in 20 neuroblastoma was 70% and the expression rate of MDR1 was 60%, but P>0.05,the result doesn't have statistically significance.Conclusion: 1. HA117 proteins in neuroblastoma is extensive distribution, prompting the neuroblastoma having multiple drug resistance. 2. HA117 was with significantly higher in lymph node and distant metastasis than without lymph node metastasis. 3.Tumours pathologic stage mayn't have relationship.