[Preparation of antibodies against a novel leucine-zipper protein, Ecp].
The aim of this research is to prepare high quality polyclonal antibodies against Ecp, a recently identified leucine-zipper protein. The full length cDNA of ecp was amplified by PCR, cloned into pGEX-4T-1(His)6 and transformed into E. coli DH 5alpha. After induction with IPTG, the GST-Ecp fusion protein from the lysate was bound to glutathione-Sepharose 4B and digested with thrombin. The released Ecp protein was further purified through Ni-NTA affinity chromatography to homogeneity. A rabbit was immunized with the purified Ecp, and the antibody generated against Ecp was purified by affinity chromatography. The results of the Western blot showed that Ecp is present in various development stages of Drosophila melanogaster, from larvae to adult.
Wang
Dao-Yong
DY
Genetics Research Center, Department of Genetics and Developmental Biology, Southeast University, Nanjing 210009, China.
Liu
Li
L
Xie
Wei
W
D000818:Animals; D000906:Antibodies / Q000276:immunology* / Q000302:isolation & purification* / Q000378:metabolism; D015153:Blotting, Western; D002846:Chromatography, Affinity; D029721:Drosophila Proteins / Q000235:genetics / Q000276:immunology*; D004331:Drosophila melanogaster / Q000254:growth & development / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D040061:Eukaryotic Initiation Factor-5; D018507:Gene Expression Regulation, Developmental / Q000235:genetics / Q000502:physiology; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism*; D010448:Peptide Initiation Factors / Q000235:genetics / Q000276:immunology*; D016133:Polymerase Chain Reaction; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism*
D000906:Antibodies; D029721:Drosophila Proteins; D040061:Eukaryotic Initiation Factor-5; C520428:Krasavietz protein, Drosophila; D010448:Peptide Initiation Factors; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins