Expression of feline immunodeficiency virusgag gene inEscherichia coli

Furuya, Tetsuya; Hasegawa, Akira; Saitoh, Masako; Miyazawa, Takayuki; Tohya, Yukinobu; Hayami, Masanori; Takahashi, Eiji; Miki, Keizaburo; Mikami, Takeshi

doi:10.1007/bf01317200

Cited by 4 publications

(4 citation statements)

References 22 publications

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“…The sections were preincubated for 1 h in phosphate-buffered saline with 0.1% Triton-X (PBS-T) containing 1% normal goat serum to prevent background staining, washed in PBS-T, and incubated in a freshly prepared solution of 0.5% hydrogen peroxide in absolute methanol for 30 min for inhibition of endogenous peroxidase. The sections were then reacted in a humidified chamber for 24 h with a rabbit anti-FIV Gag serum diluted at 1:3,200 (12,49). After the mixture was washed in PBS-T, biotinylated goat anti-rabbit immunoglobulin G serum (diluted at 1:800 in PBS-T [Dako A/S, Glostrup, Denmark]) was added, and the mixture was incubated for a further 30 min.…”

Section: Construction Of Mutantsmentioning

confidence: 99%

Roles of the auxiliary genes and AP-1 binding site in the long terminal repeat of feline immunodeficiency virus in the early stage of infection in cats

Inoshima

Kohmoto

Ikeda

et al. 1996

J Virol

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To examine the roles of auxiliary genes and the AP-1 binding site in the long terminal repeat of feline immunodeficiency virus (FIV) in vivo, three mutant viruses, which are defective in the vif gene (⌬vif), ORF-A gene (⌬ORF-A), and AP-1 binding site (⌬AP-1), and wild-type virus as a positive control were separately inoculated into three specific-pathogen-free cats. These cats were assessed by measuring the number of proviral DNA copies in peripheral blood mononuclear cells (PBMCs), the CD4/CD8 ratio and antibody responses to FIV for 16 weeks and then examining histological changes at necropsy. Although viral DNAs were detected in PBMCs from all 12 cats to various degrees until 16 weeks postinoculation, no virus was recovered from PBMCs of cats infected with ⌬vif virus during the observation period. However, a very weak antibody response was induced in one cat infected with the ⌬vif virus. In contrast, despite the successful recovery of virus from both groups of cats infected with ⌬ORF-A and ⌬AP-1 virus, antibody responses and decrease in the CD4/CD8 ratio in the groups were milder than those in cats infected with wild-type virus. Furthermore, the numbers of proviral DNA copies in PBMCs from the two groups were not able to reach the level in cats infected with wild-type virus during the observation period. From these results, we conclude that these mutant viruses are still infectious for cats but failed in efficient viral replication and suggest that these auxiliary genes and enhancer element are important or essential to full viral replication kinetics and presumably to full pathogenicity during the early stage of infection in vivo.

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Section: Construction Of Mutantsmentioning

confidence: 99%

Roles of the auxiliary genes and AP-1 binding site in the long terminal repeat of feline immunodeficiency virus in the early stage of infection in cats

Inoshima

Kohmoto

Ikeda

et al. 1996

J Virol

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“…After cocultivation of the Petaluma-and KYO-1-inoculated feline brain cells with MYA-1 cells, the culture supernatants of the cocultured MYA-1 cells contained high levels of RT activities and the specificity for the FIV infection was confirmed by the indirect immunofluorescence (IF) assay using a rabbit antiserum against FIV gag protein expressed in Escherichia coli [12] and an anti-FIV cat serum (data not shown). Further, RT activities became detectable in the supernatants of MYA-1 cells cocultured with the TM 1-inoculated brain cell cultures from the day 12 after cocultivation, indicating that the TM1 strain has much less replicative ability in the brain cells (Fig.…”

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confidence: 93%

Replicative difference in early-passage feline brain cells among feline immunodeficiency virus isolates

Kawaguchi

Maeda

Tohya

et al. 1992

Archives of Virology

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The susceptibility of early-passage feline brain cells and Crandell feline kidney (CRFK) cells to infection with three isolates of feline immunodeficiency virus (FIV) was investigated. The Petaluma strain of FIV could well infect both the feline brain cells and CRFK cells. The KYO-1 strain could well infect the feline brain cells but the replication in CRFK cells was demonstrated only by coculturing fresh feline T-lymphoblastoid cells with the infected cells. On the other hand, the TM1 strain could infect the feline brain cells but not CRFK cells. Moreover, the replicative ability of the TM1 strain in the feline brain cells was much less than the KYO-1 and Petaluma strains. These results indicate that biological differences can be detected among the FIV isolates.

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“…Among these ORFs, gag encodes the viral core and capsid proteins. Recently, we developed an enzyme-linked immunosorbent assay (ELISA) system using the expressed gag proteins in Escheriehia coli to detect anti-gag antibodies, and the degree of ELISA value (EV) found to correspond to the intensity of p26 band of the major core protein detected by Western blotting analysis [3]. In this report, we investigated serological responses to the gag protein in cats experimentally and naturally infected with FIV using the ELISA and a radioimmunoprecipitation assay (RIPA).…”

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confidence: 99%

“…The gag antigen was prepared from E. coli HB 101 expressing FIV gag protein as described previously [3], and control antigen was obtained from intact E. coli by the same manner as prepared the gag antigen. The methods of the immune and enzymatic reactions were described previously [3]. After enzymatic reaction, optical density at 415 nm was read for the both types of plates.…”

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confidence: 99%

Detection of anti-gag antibodies of feline immunodeficiency virus in cat sera by enzyme-linked immunosorbent assay

Furuya

Hasegawa²,

Miyazawa

et al. 1992

Archives of Virology

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Using gag protein of feline immunodeficiency virus (FIV) expressed in Escherichia coli, an enzyme-linked immunosorbent assay (ELISA) system was developed for detection of antibodies to FIV gag protein in cat sera. With serum samples from cats experimentally infected with several strains and an infectious molecular clone of FIV, increases of the antibody titers to FIV gag protein were observed in all cases by the ELISA at early stage of infection. When we examined a total of 415 field cat sera which were previously tested by an indirect immunofluorescence assay (IFA), 9 (12.9%) out of 70 IFA positive sera were judged as negative by the ELISA. However, all 3 serum samples tested among the 9 IFA positive sera had antibodies to gp130 but not to p26 by a radioimmunoprecipitation assay. The results indicated that some IFA positive sera did not have antibodies to the p26 though they have antibodies to other proteins specific for FIV.

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Expression of feline immunodeficiency virusgag gene inEscherichia coli

Cited by 4 publications

References 22 publications

Roles of the auxiliary genes and AP-1 binding site in the long terminal repeat of feline immunodeficiency virus in the early stage of infection in cats

Roles of the auxiliary genes and AP-1 binding site in the long terminal repeat of feline immunodeficiency virus in the early stage of infection in cats

Replicative difference in early-passage feline brain cells among feline immunodeficiency virus isolates

Detection of anti-gag antibodies of feline immunodeficiency virus in cat sera by enzyme-linked immunosorbent assay

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