Analysis of cellular immune responses in the peripheral blood of mice using real-time RT-PCR

Hempel, D; Smith, Karen; Claussen, Kirsten A; Perricone, Michael A.

doi:10.1016/s0022-1759(01)00502-6

Cited by 27 publications

(14 citation statements)

References 22 publications

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“…The sequences of primers and probes designed with Primer Express Software and used for quantifying IFN-c were as follows [27]:…”

Section: Real-time Quantitative Reverse Transcriptase-polymerase Chaimentioning

confidence: 99%

Surgical excision combined with autologous whole tumor cell vaccination is an effective therapy for murine neuroblastoma

Ohashi

Kobayashi

Fang

et al. 2006

Journal of Pediatric Surgery

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“…The sequences of primers and probes designed with Primer Express Software and used for quantifying IFN-c were as follows [27]:…”

Section: Real-time Quantitative Reverse Transcriptase-polymerase Chaimentioning

confidence: 99%

Surgical excision combined with autologous whole tumor cell vaccination is an effective therapy for murine neuroblastoma

Ohashi

Kobayashi

Fang

et al. 2006

Journal of Pediatric Surgery

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“…Its potential as a clinical diagnostic assay (Bustin & Dorudi 1998) is also being realised, and its use has been reported for the identification of micrometastases or minimal residual disease in colorectal cancer (Bustin et al 1999), neuroblastoma (Cheung & Cheung 2001), prostate cancer (Gelmini et al 2001) and leukaemia (Buonamici et al 2002). It has been used to distinguish different types of lymphoma (Bijwaard et al 2001), for the analysis of cellular immune responses in the peripheral blood (Hempel et al 2002), the detection of bacterial (Goerke et al 2001) and viral (Greijer et al 2002) RNA in clinical samples and for monitoring the response of human cancer to drugs (Miyoshi et al 2002). Other clinically relevant applications include its use for the detection of gene amplification in breast cancer (Bieche et al 1999), for the analysis of tissue-specific gene expression and for cytokine mRNA profiling (Stordeur et al 2002).…”

Section: Introductionmentioning

confidence: 99%

Quantification of mRNA using real-time reverse transcription PCR (RT-PCR): trends and problems

Bustin

2002

Journal of Molecular Endocrinology

2,234

1,674

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The fluorescence-based real-time reverse transcription PCR (RT-PCR) is widely used for the quantification of steady-state mRNA levels and is a critical tool for basic research, molecular medicine and biotechnology. Assays are easy to perform, capable of high throughput, and can combine high sensitivity with reliable specificity. The technology is evolving rapidly with the introduction of new enzymes, chemistries and instrumentation. However, while real-time RT-PCR addresses many of the difficulties inherent in conventional RT-PCR, it has become increasingly clear that it engenders new problems that require urgent attention. Therefore, in addition to providing a snapshot of the state-of-the-art in real-time RT-PCR, this review has an additional aim: it will describe and discuss critically some of the problems associated with interpreting results that are numerical and lend themselves to statistical analysis, yet whose accuracy is significantly affected by reagent and operator variability.

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“…Thus, we hypothesized that HSV infection might induce prolonged expression of a broad range of chemokines at sites of acute and latent infection. Real-time quantitative RT-PCR methods have facilitated studies of immune cell RNA expression in mouse models [22,23]. We report here the use of real-time RT-PCR to monitor RNA expression of selected chemokine receptors and their chemokine ligands during HSV infection of mouse corneal and TG tissue.…”

Section: Introductionmentioning

confidence: 99%

Persistent expression of chemokine and chemokine receptor RNAs at primary and latent sites of herpes simplex virus 1 infection

Cook

Kramer

Walker

et al. 2004

Virol J

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Inflammatory cytokines and infiltrating T cells are readily detected in herpes simplex virus (HSV) infected mouse cornea and trigeminal ganglia (TG) during the acute phase of infection, and certain cytokines continue to be expressed at lower levels in infected TG during the subsequent latent phase. Recent results have shown that HSV infection activates Toll-like receptor signaling. Thus, we hypothesized that chemokines may be broadly expressed at both primary sites and latent sites of HSV infection for prolonged periods of time. Real-time reverse transcriptase-polymrease chain reaction (RT-PCR) to quantify expression levels of transcripts encoding chemokines and their receptors in cornea and TG following corneal infection. RNAs encoding the inflammatory-type chemokine receptors CCR1, CCR2, CCR5, and CXCR3, which are highly expressed on activated T cells, macrophages and most immature dendritic cells (DC), and the more broadly expressed CCR7, were highly expressed and strongly induced in infected cornea and TG at 3 and 10 days postinfection (dpi). Elevated levels of these RNAs persisted in both cornea and TG during the latent phase at 30 dpi. RNAs for the broadly expressed CXCR4 receptor was induced at 30 dpi but less so at 3 and 10 dpi in both cornea and TG. Transcripts for CCR3 and CCR6, receptors that are not highly expressed on activated T cells or macrophages, also appeared to be induced during acute and latent phases; however, their very low expression levels were near the limit of our detection. RNAs encoding the CCR1 and CCR5 chemokine ligands MIP-1α, MIP-1β and RANTES, and the CCR2 ligand MCP-1 were also strongly induced and persisted in cornea and TG during the latent phase. These and other recent results argue that HSV antigens or DNA can stimulate expression of chemokines, perhaps through activation of Toll-like receptors, for long periods of time at both primary and latent sites of HSV infection. These chemokines recruit activated T cells and other immune cells, including DC, that express chemokine receptors to primary and secondary sites of infection. Prolonged activation of chemokine expression could provide mechanistic explanations for certain aspects of HSV biology and pathogenesis.

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Analysis of cellular immune responses in the peripheral blood of mice using real-time RT-PCR

Cited by 27 publications

References 22 publications

Surgical excision combined with autologous whole tumor cell vaccination is an effective therapy for murine neuroblastoma

Surgical excision combined with autologous whole tumor cell vaccination is an effective therapy for murine neuroblastoma

Quantification of mRNA using real-time reverse transcription PCR (RT-PCR): trends and problems

Persistent expression of chemokine and chemokine receptor RNAs at primary and latent sites of herpes simplex virus 1 infection

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