For fast, one-step qRT-PCR using sequence-specific probes for gene expression analysis
Faster results with time savings of up to 60%
Sensitive detection of even low copy numbers
Accurate detection of a wide range of template amounts
Optimized, ready-to-use master mix for fast cycling
One cycling protocol for all standard and fast cyclers
QuantiFast Probe RT-PCR Kits deliver fast and sensitive real-time one-step RT-PCR quantification of RNA targets using probe-based detection. Q-bond technology and an optimized master mix enable shorter real-time RT-PCR run times, not only on fast cyclers with short ramping times, but also on standard cyclers. The combination of a hot start and a unique PCR buffer system in the ready-to-use master mix ensures highly sensitive qPCR on any real-time cycler without the need for optimization. The kits are also supplied with an optimized RT mix for efficient cDNA synthesis in only 10 minutes. Two kit formats are available: the QuantiFast Probe RT-PCR Kit for cyclers that require ROX dye for fluorescence normalization, and the QuantiFast Probe RT-PCR +ROX Vial Kit for all other cyclers. For convenience, the master mix can be stored at 2–8°C.
For 2000 x 25 µl reactions: 25 ml 2x QuantiFast Probe RT-PCR Master Mix (without ROX dye), 1.05 ml ROX Dye Solution, 0.5 ml QuantiFast RT Mix, 20 ml RNase-Free Water
Significantly reduced RT-PCR times.
QuantiFast Probe Kits reduce total RT-PCR run time by up to 60% in real-time one-step RT-PCR on standard cyclers (40 PCR cycles carried out; comparison with a standard QIAGEN real-time PCR kit). L: LightCycler 2.0; A1: ABI PRISM 7900; A2: Applied Biosystems 7500; A3: ABI PRISM 7000.
Sensitive one-step RT-PCR.
Reactions were run in duplicate using 10-fold dilutions of human leukocyte RNA (100 ng to 0.1 ng) and a Primer Express designed TaqMan assay for IL12RBI (a type I transmembrane protein) on the Applied Biosystems 7500 Fast System. [A] The QuantiFast Kit showed greater sensitivity than [B] the kit from Supplier I (which was used according to the standard-cycling protocol), providing lower CT values and transcript quantification from 0.1 ng RNA.
Comparable dynamic range.
Ubiquitin expression in human leukocytes was analyzed using a Primer Express designed TaqMan assay. Two-step and one-step qPCR reactions were run in duplicate on the ABI PRISM 7900. [A] 10-fold cDNA dilutions (100 ng to 10 pg) were analyzed using the QuantiFast Probe PCR Kit. [B] 10-fold RNA dilutions (100 ng to 10 pg) were analyzed using the QuantiFast Probe RT-PCR Kit.
Wide dynamic range and high sensitivity.
Triplicate reactions were performed using 10-fold dilutions of human leukocyte RNA and a TaqMan assay for β-actin. The QuantiFast Probe RT-PCR Kit provided accurate gene expression analysis from 100 ng down to 1 pg of RNA template.
QuantiFast Probe RT-PCR Kits deliver highly sensitive results, outperforming other real-time RT-PCR kits (see figure "Sensitive One-step RT-PCR"). RT-PCR run times are reduced by up to 60% (see figure "Significantly reduced PCR times"), allowing you to achieve fast PCR results without compromising on RT-PCR performance (see figure "Faster results without compromising sensitivity"). You can also greatly increase your sample throughput or efficiently share a cycler with other users. With QuantiFast Probe RT-PCR Kits, you follow the same procedure whether you work with a standard or fast cycler.
QuantiFast Probe RT-PCR Kits deliver highly sensitive and rapid results over a wide dynamic range on both standard and fast cyclers. The kits are designed for use with all types of sequence-specific probes, including hydrolysis probe detection (e.g., TaqMan and other dual-labeled probes) and FRET probes. The optimized QuantiFast RT Mix enables cDNA synthesis in just 10 minutes and a specially developed fast RT-PCR buffer contains the novel additive Q-Bond, which significantly reduces denaturation, annealing, and extension times (see figure "Fast primer annealing"). The RT-PCR buffer also contains a balanced combination of K+ and NH4+ ions, which promote specific primer annealing and enable high RT-PCR specificity and sensitivity (see figure "Specific primer annealing"). In addition, HotStarTaq Plus DNA Polymerase requires only 5 minutes at 95°C for activation and provides a stringent hot start, preventing the formation of nonspecific products.
Components of 2x QuantiFast Probe RT-PCR Kit*
Features and benefits
HotStarTaq Plus DNA Polymerase
5 min activation at 95ºC
Set up of qPCR reactions at room temperature
QuantiFast Probe RT-PCR Buffer
Balanced combination of NH4+ and K+ ions
Specific primer annealing ensures reliable PCR results
Unique Q-Bond additive
Faster PCR run times, enabling faster results and more reactions per day
Normalizes fluorescent signals on Applied Biosystems and, optionally, Agilent instruments
Precise quantification on cyclers that require ROX dye. Does not interfere with PCR on any real-time cycler
QuantiFast RT Mix
Separate solution added during PCR setup
Fast cDNA synthesis in just 10 minutes
* Also contains dNTP mix (dATP, dCTP, dGTP, and dTTP). † ROX dye is either present in the master mix or as a separate solution.
QuantiFast Probe RT-PCR Kits contain ready-to-use master mixes that eliminate the need for optimization of reaction and cycling conditions. Simply add template RNA, primers, and probe to the master mix and follow the protocol in the handbook to get fast and reliable results on any real-time cycler. Kits are available with or without ROX passive reference dye in the master mix, enabling use on virtually any real-time cycler (see table). Due to the optimized ROX concentrations, detection of even low copy numbers is achieved through automatic data analysis.
Choosing the right QuantiFast Probe RT-PCR Kit
Supplied in master mix
QuantiFast Probe RT-PCR Kit
All cyclers from Applied Biosystems except Applied Biosystems 7500
Supplied in separate tube
QuantiFast Probe RT-PCR +ROX Vial Kit
Applied Biosystems 7500 and cyclers from Bio-Rad, Cepheid, Eppendorf, QIAGEN, Roche, Agilent, and other suppliers
QuantiFast Probe RT-PCR Kits can be used for probe-based gene expression analysis of RNA targets on any real-time cycler. This includes instruments from Applied Biosystems, Bio-Rad, Cepheid, Eppendorf, Roche, and Agilent. For the Rotor-Gene Q and other Rotor-Gene cyclers, we recommend using the Rotor-Gene Probe RT-PCR Kit, which has been specially developed for fast cycling on these instruments.
Probe-based, real-time RT-PCR
Real-time one-step RT-PCR
Real-time or endpoint
Single or multiplex
SYBR Green I or sequence-specific probes
All real-time cyclers (e.g. LC, RG, ABI)
With or without ROX
Available with ROX in master mix and with ROX as a separate vial
For 100 x 50 µl reactions: QIAGEN OneStep RT-PCR Enzyme Mix (1 x 200 µl), 5x QIAGEN OneStep RT-PCR Buffer (1 x 1 ml), dNTP Mix (1 x 200 µl, 10 mM each), 5x Q-Solution (1 x 2 ml), RNase-Free Water (2 x 1.9 ml)