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- Production and Characterization of Monoclonal Antibodies to Human Immunodeficiency Virus Type 1 gp120 Envelope Glycoprotein
- Choi, Eui Yul , Ryu, Ji Yoon , Lee, Yoon , Ha, Sung Gil , Chung, So Young , Park, Sang Yeol , Nham, Sang Uk , Lee, Young Ik , Park, Jin Seu
- J. Microbiol. 1998;36(1):59-65.
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Abstract
- Monoclonal antibodies to human immunodeficiency virus type 1 envelope glycoprotein gp 120(HIV-1 gp120) were produced and characterized. For immunogen recombinant gp120 polypeptide expressed in bacteria was prepared and injected into mice. From two fusion experiments, twenty hybridomas secreting monoclonal antibodies against the recombinant gp120 were initially screened by immunodot blot analysis. Among the antibodies, 15 of them showed strong reactivities with the recombinant protein expressed in bacteria in Western blot and thus it was tested if these could react with the recombinant protein expressed in insect cells. All of the 15 antibodies immunostained the protein band with varing degrees of reactivities. Next, we tested whether the antibodies recognize authentic gp120 protein expressed in mammalian cells. COS-1 cells were tranfected with the cDNA encoding gp120 protein, and the transiently ecpressed protein were analyzed with the mAbs by Western blot analysis and immunofluorescence microscopy. Six of the monoclonal antibodies reacted with the protein band of authentic gp120 expressed in mammalian cells in the Western blot, and five stained the cell periphery of the transfected COS-1 cells in immunofluorescence. The mAbs described in this study should prove to be useful tools for the biochemical, immunological and structural analysis of HIV-1 gp120 envelope glycoprotein.
- Expression of Human Cytomegalovirus Immediate Early US3 Gene in Human Fibroblast Cells
- Gyu-Cheol Lee , Chong-Kyo Lee , Jin-Hyun Ahn , Chan-Hee Lee
- J. Microbiol. 2000;38(1):24-30.
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Abstract
- US3 gene is a member of the human cytomegalovirus (HCMV) immediate early gene. Although the precise functions of the US3 gene in HCMV replication and pathogenesis are not known, it has been reported to play a role in inhibiting major histocompatibility class I antigen presentation. For further knowledge of US3 gene expression, rabbit polyclonal antiserum of the US3 gene product was used for indirect immunofluorescence assay. In permissive human foreskin fibroblast (HFF) cells, US3 gene expression was detectable as crescent or half-moon shape in the perinuclear region at immediate early times after virus infection. HFF cells infected with mutant HCMV lacking US3 open reading frames were negative for US3 immunofluorescence assay. Double immunofluorescence assay using monoclonal antibody to gamma adaptin (specific for the Golgi complex) and rabbit anti-US3 antiserum revealed that US3 gene product could be localized to the Golgi complex. At later time after HCMV infection, US3 gene products were detected as globular aggregates in the cytosol. These aggregates were positive for gamma adaptin and stained with preimmune serum, suggesting a nonspecific reaction to the Golgi complex. Northern blot analysis revealed that transcription of US3 was observed only during immediate early times after virus infection (until 6 h postinfection). Therefore US3 gene expression appears to be confined to immediate early time and its gene products are localized to the Golgi complex as cres-cent shaped forms in the perinuclear cytoplasm.
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