Infectivity of Rous Sarcoma Cell DNA: Comparison of Two Techniques of Transfection Assay

Hillova, Jana; Hill, M.; Goubin, Gérard; Dantchev, Dimitre

doi:10.1159/000149935

Cited by 12 publications

(9 citation statements)

References 13 publications

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“…DNA from recombinant colonies was amplified by Qiagen plasmid maxi kit and transfected into ecotropic packaging cells (PE501) by using a calcium phosphate method (16). PE501 viral supernatant was used to infect PG13 cells, a primatespecific packaging line.…”

Section: Cd83 Retrovirus Construction and Generation Of Transfected Cmentioning

confidence: 99%

CD83 Is a Sialic Acid-Binding Ig-Like Lectin (Siglec) Adhesion Receptor that Binds Monocytes and a Subset of Activated CD8+ T Cells

Scholler¹,

Hayden-Ledbetter

Hellström³

et al. 2001

The Journal of Immunology

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To help determine CD83 function, a cDNA encoding a soluble protein containing the CD83 extracellular domain was fused with a mutated human IgG1 constant region (CD83Ig) and expressed by stable transfection of Chinese hamster ovary cells. Purified CD83Ig bound to peripheral blood monocytes and a subset of activated CD3+CD8+ lymphocytes but did not bind to FcR. Monocytes that had adhered to plastic lost their ability to bind to CD83Ig after 90 min of in vitro incubation. CD83Ig bound to two of five T cell lines tested, HPB-ALL and Jurkat. The binding to HPB-ALL cells significantly increased when they were grown at a low pH (pH 6.5), whereas binding to Jurkat cells increased after apoptosis was induced with anti-Fas mAb. B cell and monocytic lines did not bind CD83Ig and neither did CD56+ NK cells or granulocytes. Full-length CD83 expressed by a transfected carcinoma line mediated CD83-dependent adhesion to HPB-ALL cells. CD83Ig immunoprecipitated and immunoblotted a 72-kDa protein from HPB-ALL cells. Binding of CD83Ig to HPB-ALL cells was eliminated by neuraminidase treatment of the cells. We conclude that CD83 is an adhesion receptor with a counterreceptor expressed on monocytes and a subset of activated or stressed T lymphocytes, and that interaction between CD83 and its counterreceptor is dependent upon the state of glycosylation of a 72-kDa counterreceptor by sialic acid residues. In view of the selectivity of the expression of CD83 and its ligand, we postulate that the interaction between the two plays an important role in the induction and regulation of immune responses.

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Section: Cd83 Retrovirus Construction and Generation Of Transfected Cmentioning

confidence: 99%

CD83 Is a Sialic Acid-Binding Ig-Like Lectin (Siglec) Adhesion Receptor that Binds Monocytes and a Subset of Activated CD8+ T Cells

Scholler¹,

Hayden-Ledbetter

Hellström³

et al. 2001

The Journal of Immunology

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“…It has been shown previously that this proce dure has the same efficiency as the calcium phosphate technique when transfected cells are propagated under liquid medium [18]. In the case of transfections under agar overlay, however, the DEAE-dextran technique was abandoned since we found it to be far less efficient than calcium phosphate.…”

mentioning

confidence: 86%

mentioning

confidence: 99%

“…The extraction proce dure was reported elsewhere [18], except that the cell lysate was digested overnight with 50 ¡ag/ml proteinase K (Boehringer Mann heim France SA) instead of pronase. The transfection technique using calcium phosphate-DNA coprecipitation [19] has also been described [18], The assay was modified as follows. About 1.2x10° chicken or tur key cells in 5 ml of modified [20] Eagle medium with 10% tryptose phosphate broth (TPB: Difco, Detroit, Mich.) and 5 or 10% heated calf serum, respectively, were seeded into 60-mm diameter Falcon Petri dishes.…”

mentioning

confidence: 99%

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Two-Hit Kinetics of Focus Formation in Cells Transfected with Rous Sarcoma Provirus

Hillova¹,

Hill²

1980

Intervirology

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DNA was purified from an established line of SR-E-transformed quail cells and was assayed for transfection in turkey cells. Transfection events were determined from foci of transformed cells grown under agar overlay which was used to prevent the spread of the progeny virus. It was found that transfection with Rous sarcoma provirus follows two-hit kinetics, indicating that at least two (unlinked) proviruses are required to give rise to a focus of transformed cells.

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“…The infectivity of DNA was tested under condi tions employing the calcium phosphate technique of [28] as described previously [29], DNA samples were diluted 10 times serially in HEPESbuflered saline, and the final concentration of DNA was adjusted to 10 ¡Jg/ml by the addition of carrier rat thymus DNA toeach dilution. After the DNA treat ment, the cells were subcultured 3 times during 3 weeks and examined both for foci of transformed cells and by reverse transcriptase assay [17] for nontransforming virus.…”

Section: Dna Infectivity Assaymentioning

confidence: 99%

Tandem Integration of Avian Retroviruses in Double-Infected Cells

Carloni¹,

N²,

Hillova³

et al. 1980

Intervirology

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Chicken embryo fibroblasts were chronically infected by both RAV-1 and SR-E, and established SR-E-transformed quail cells were superinfected with RAV-1. About 75 and 50% of double-infected and superinfected cells, respectively, produced both parental viruses. The DNA purified from these cells contained infectious provirus of both parents and no detectable recombinant provirus. In assays using agar overlay, about 5% of the transformed foci that were induced by the DNA of double-infected, but not superinfected, cells were found to contain the progeny of both parents. This observation suggests that in double infections with avian retroviruses, DNA of both parent viruses may integrate into the host genome either in tandem or in close proximity.

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Infectivity of Rous Sarcoma Cell DNA: Comparison of Two Techniques of Transfection Assay

Cited by 12 publications

References 13 publications

CD83 Is a Sialic Acid-Binding Ig-Like Lectin (Siglec) Adhesion Receptor that Binds Monocytes and a Subset of Activated CD8+ T Cells

CD83 Is a Sialic Acid-Binding Ig-Like Lectin (Siglec) Adhesion Receptor that Binds Monocytes and a Subset of Activated CD8+ T Cells

Two-Hit Kinetics of Focus Formation in Cells Transfected with Rous Sarcoma Provirus

Tandem Integration of Avian Retroviruses in Double-Infected Cells

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