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Real Time RT-PCR

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.
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Related Experiment Video

Updated: May 20, 2025

Simple Bulk Readout of Digital Nucleic Acid Quantification Assays

Simple Bulk Readout of Digital Nucleic Acid Quantification Assays

Published on: September 24, 2015

Chip-based digital PCR with large-field one-shot imaging for high-sensitivity nucleic acid quantification.

Jinrong Shen¹, Jingxing Fan¹, Zhehao Zhao²

¹State Key Laboratory of ASIC and System, Fudan University, Shanghai, 200433, China; School of Microelectronics, Fudan University, Shanghai, 200433, China.

Biosensors & Bioelectronics

|March 25, 2025

Summary

This summary is machine-generated.

We developed a chip-based digital polymerase chain reaction (cdPCR) device with a novel large-field imaging system. This system enables rapid, one-shot image acquisition, improving absolute quantification in dPCR analysis.

Keywords:

Absolute quantification Chip-based digital PCR Fluorescence image analysis High-uniformity illumination system Lung cancer DNA One-shot imaging

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Published on: August 20, 2018

Area of Science:

Biotechnology
Analytical Chemistry
Medical Diagnostics

Background:

Digital polymerase chain reaction (dPCR) offers high sensitivity for absolute quantification.
Traditional dPCR imaging systems face limitations in speed and accuracy due to image stitching.

Purpose of the Study:

To develop an enhanced chip-based dPCR (cdPCR) device with a large-field imaging system.
To overcome limitations of traditional dPCR imaging, improving absolute detection limits and accuracy.

Main Methods:

Designed a cdPCR device with 21,384 microreaction chambers.
Developed a large-field fluorescence imaging system (28 mm × 20 mm) for one-shot image acquisition.
Implemented Köhler illumination and image processing modules for enhanced image quality and precision.

Main Results:

Achieved one-shot fluorescence image acquisition of the entire cdPCR chip in 1 second.
Demonstrated 90% illumination uniformity for high-quality imaging.
Validated the system with lung cancer DNA, showing excellent correlation (R² > 0.9997) between measured and theoretical values.

Conclusions:

The developed system provides an automated, rapid, and accurate platform for cdPCR fluorescence image processing.
Elimination of image stitching errors enhances the precision of absolute quantification.
The system advances dPCR analysis for improved diagnostic applications.