Mouse Anti Human 6XHis peptide, linear Hybridoma [Q6B22]

This study reports synthesis of Ni-nitrilotriacetic acid (Ni-NTA) modified carbon nanospheres containing magnetic Fe3O4 particles (C@Fe3O4), which can act as a general tool to separate and purify histidine-tagged fetidin. In this experiment, C nanospheres are prepared from glucose using the hydrothermal process, magnetic Fe3O4 are packed in the porous C nanospheres and Ni-NTA are conjugated to C@Fe3O4. C@Fe3O4/NTA-Ni nanospheres are about 400 nm in diameter, providing the nanospheres with excellent magnetic response and dispersity. This study separates recombinant fetidin that are engineered to have six consecutive histidine residues (6xHis) by treating with C@Fe3O4/NTA-Ni magnetic nanospheres. 10 mg of C@Fe3O4/NTA-Ni nanospheres can purify up to 2.103 mg of high-purity 6xHis-tagged fetidin from 10 ml of crude Escherichia coli lysates. Because of high performance,

Production of structural proteins from foot-and-mouth disease virus (FMDV) and bovine herpes virus (BHV-1) in Nicotiana benthamiana through the use of a tobacco mosaic virus-based vector (TMV-30B) has been reported previously. The development of the TMV-30B-HISc vector, a new version that adds a C-terminal histidine (His) sequence to the foreign protein expressed is described. Coding sequences from the FMDV VP1 protein and the core protein, p24, from a clade C HIV-1 isolate from Zambia were cloned into the new vector and infective RNAs were generated for each construct to inoculate N. benthamiana plants. His-tagged proteins were purified from inoculated leaves using immobilized metal affinity chromatography (IMAC) as detected by Coomassie blue staining and proteins were further characterized in Western blot assays using a commercial anti-6xHis mAb and specific polyclonal antisera for each protein. While yields obtained for the VP1-His protein after purification were similar to those in crude extracts obtained with the previous TMV-VP1 vector, p24-His yields were 10-15 times higher than those of VP1-His. Twenty-five grams of TMV-p24-HISc inoculated leaves were processed to obtain 2.5 mg of isolated p24-His and the recombinant protein was inoculated in rabbits to test immunogenicity and antigenic integrity of the plant-produced p24-His. Animals developed a strong and specific Immoral response to the p24-His after the first booster and immune sera was able to recognize the native p24 from a different clade expressed on the surface of the HIV-1 chronically infected HUT78/ARV T-cell line. Importantly, the recombinant p24-His proved its efficiency by confirming the serology of 117 samples previously tested by two rapid HIV-1 tests, thus representing an excellent alternative for production of highly specific diagnostic reagents for HIV endemic regions in the developing world. (C) 2004 Elsevier B.V. All rights reserved.