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Related Concept Videos

Real Time RT-PCR02:57

Real Time RT-PCR

Real-time reverse transcription-polymerase chain reaction, or Real-time RT-PCR, is an analytical tool used to determine the expression level of target genes. The method involves converting mRNA to complementary DNA with the help of an enzyme known as reverse transcriptase, followed by the PCR amplification of the cDNA. These two processes can be performed simultaneously in a single tube or separately as a two-step reaction.
The real-time quantification of the number of amplified products is...

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Preparation of Peripheral Blood Mononuclear Cell Pellets and Plasma from a Single Blood Draw at Clinical Trial Sites for Biomarker Analysis
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Quantifying a bystander response following microbeam irradiation using single-cell RT-PCR analyses.

Brian Ponnaiya1, Gloria Jenkins-Baker, Gerhard Randers-Pherson

  • 1Center for Radiological Research, Columbia University, New York, NY 10533, USA. bp156@columbia.edu

Experimental Hematology
|March 24, 2007
PubMed
Summary

Ionizing radiation affects neighboring cells, not just directly hit cells. This study used a microbeam to show that non-hit cells exhibit bystander effects, with altered gene expression following radiation exposure.

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Area of Science:

  • Radiation biology
  • Cellular and molecular biology
  • Genetics

Background:

  • The effects of ionizing radiation extend beyond directly damaged cells, impacting neighboring cells through bystander effects.
  • Understanding these bystander effects is crucial for accurate radiation risk assessment.

Purpose of the Study:

  • To investigate the cellular and molecular responses of individual cells to targeted irradiation using a microbeam facility.
  • To quantify the bystander effect in non-hit cells within a population exposed to precisely delivered radiation.

Main Methods:

  • Utilized a microbeam facility to deliver precise numbers of alpha-particles to specific locations (nucleus or cytoplasm) of individual human fibroblasts.
  • Quantified alterations in CDKN1a (p21/Cip1/WAF1) mRNA levels in single cells using semiquantitative RT-PCR.
  • Examined responses in both directly hit cells and non-hit bystander cells within the same population.

Main Results:

  • Single targeted cells showed increased CDKN1a mRNA levels, consistent with known radiation responses.
  • Non-hit bystander cells also exhibited elevated CDKN1a mRNA levels, albeit to a lesser extent than hit cells.
  • Demonstrated the ability to analyze individual cell responses in both targeted and bystander cells simultaneously.

Conclusions:

  • The microbeam approach effectively allows for the precise irradiation of specific cells and the subsequent quantification of responses in individual targeted and bystander cells.
  • This methodology provides a powerful tool for studying the complex mechanisms underlying radiation-induced bystander effects.