Multiplex qPCR

Traditional Real-Time PCR assays amplify a single target in each well. Multiplex qPCR amplifies more than one target in a single well using multiple fluorophores. The ability to analyze more than one target per well allows users to generate more data points and decrease reagent costs.

Multiplex qPCR can be used for a variety of applications such as gene expression, viral load, Single Nucleotide Polymorphisms (SNP) genotyping, and microbial detection and quantification. The IntelliQube® has five channels available for excitation and detection, providing a flexible instrument for multiplex applications.

Available Excitation and Emission Channels on the IntelliQube.

LEDs (470-627 nm)
Dye Channel
1
2
3
4
5
Excitation Filter (nm)
480
530
548
580
620
Emission Filter (nm)
510
565
580
625
705
Common Dyes
FAM™
JOE™
VIC®/HEX™
Cal Fluor® Orange 560
NED™
TAMRA™
Cal Fluor Red 590
ROX™
Cal Fluor Red 610
Texas Red®
Cy5®
Quasar® 670

FAM, JOE, VIC, HEX, NED, TAMRA and ROX are trademarks of Life Technologies Corporation and its subsidiaries. VIC and Texas Red are registered trademarks of Life Technologies Corporation and its subsidiaries. Cy5 is a registered trademark of GE Healthcare Bio-Sciences Corp. Cal Fluor and Quasar are registered trademarks of Biosearch Technologies, Inc.

In a recent study, two pharmacologically relevant SNP CYP2C9*2 and VKORC1 were analyzed in a single well using the IntelliQube System (figure one).

Figure 1

This study demonstrated the ability for the IntelliQube to successfully multiplex two SNP assays in a single well. The combination of achieving two results per reaction well has a dramatic impact on overall throughput (Table 1).

Standard
80
1
6
768
4,608
Fast
35
1
14
768
10,752
Fast Multiplex
35
2
14
1,536
21,504

Table 1: Determination of the number of data points achievable in an eight-hour day.

Click here to download the complete study!.