title: [Construction,  expression and characterization of the fusion gene of super-antigen SEA and single chain Fv of the ND-1 monoclonal antibody against human colorectal cancer].
abstract: OBJECTIVE
  To construct  and express the recombinant ND-1-scFv/SEA,  a fusion protein of superantigen (staphylococcal enterotoxinA, SEA)  and single-chain variable fragment of monoclonal antibody ND-1 against human clolorectal carcinoma,  and to enhance the targeted killing effect of SEA.
  
  
  METHODS
  The expression of the fusion protein was induced in E.coli M15 by IPTG. Ni-NTA resin affinity chromatography was used to separate and purify the expressed product. The specific binding activity of the purified ND-1-scFv/SEA protein was examined by indirect immunofluorescence assay and the targeted-cytotoxicity was determined using MTT assay.
  
  
  RESULTS
  The expressing vector of fusion gene ND-1scFv/SEA was constructed successfully. ND-1-scFv/SEA protein retained a high binding affinity to antigen-positive human colorectal cancer cell CCL-187 and had a stronger capability to activate PBMC and kill the target cells compared to SEA alone,  with a killing rate of 91% at 4  μg/mL.
  
  
  CONCLUSIONS
  ND-1-scFv/SEA fusion protein could specifically target colorectal cancer cell,  enhance the activity of kill tumor cell and has potential applications in the targeted therapy of colorectal cancer.
authors.affiliation: Department of Cell Biology,  Key Labaratory of Cell Biology,  Ministry of  Public Health,  and Key Labaratory of Medical Cell Biology,  Ministry of Education, China  Medical University,   Shenyang  110001,  China.
authors.forename: Hang
authors.initials: H
authors.lastname: Chen
authors.forename: Li
authors.initials: L
authors.lastname: Li
authors.forename: Jin
authors.initials: J
authors.lastname: Fang
mesh_terms: D000915:Antibody Affinity / Q000276:immunology; D045744:Cell Line, Tumor; D002478:Cells, Cultured; D003001:Cloning, Molecular; D015179:Colorectal Neoplasms / Q000276:immunology* / Q000378:metabolism; D003602:Cytotoxicity, Immunologic / Q000276:immunology; D004591:Electrophoresis, Polyacrylamide Gel; D004768:Enterotoxins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000276:immunology / Q000378:metabolism; D011485:Protein Binding / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D057127:Single-Chain Antibodies / Q000235:genetics / Q000276:immunology* / Q000378:metabolism
chemical_list: D004768:Enterotoxins; C475886:Nd-1scFv; D011993:Recombinant Fusion Proteins; D057127:Single-Chain Antibodies; C026066:enterotoxin A, Staphylococcal