QuantiTect Virus Kits

For highly sensitive detection of viral RNA and/or DNA

Features

  • High sensitivity in single and multiplex assays
  • Detection of viral RNA and/or DNA in the same reaction
  • Clear detection of weak positive signals
  • Fast universal 2-step protocols
  • 5x master mix for higher sensitivity with more sample input
QuantiTect Virus Kit (1000)

Cat. No. / ID: 211015

For 1000 x 50 µl reactions: QuantiTect Virus Master Mix (contains ROX dye), QuantiTect Virus RT Mix, RNase-Free Water, QuantiTect Nucleic Acid Dilution Buffer
Reference dye
Rox in Mastermix
Rox in vial
Reactions
1000
200
50
QuantiTect Virus Kits are intended for molecular biology applications. These products are not intended for the diagnosis, prevention, or treatment of a disease.

Product Details

The QuantiTect Virus Kit is specially designed for highly sensitive detection of up to 4 viral nucleic acid targets by multiplex, real-time one-step RT-PCR using sequence-specific probes. Multiple viral RNA and/or DNA targets plus internal controls can be detected in a single tube without loss of sensitivity. The concentrated 5x master mix enables the addition of higher sample input and contains HotStarTaq Plus DNA Polymerase. The QuantiTect Virus RT Mix contains a unique formulation of Sensiscript Reverse Transcriptase that is optimized for highly sensitive detection of viral RNA. Two kit formats are available: the QuantiTect Virus Kit for cyclers that require ROX dye for fluorescence normalization and the QuantiTect Virus +ROX Vial Kit for other cyclers. For convenience, the master mix can be stored at 2–8°C.

Performance

Amplification using QuantiTect Virus Kits provides steep sigmoidal curves over a range of dilutions, even for low template amounts with high CT values (see figure "  Unambiguous determination of CT values over a wide dynamic range"). This enables accurate CT value determination for quantification of viral nucleic acids in real-time PCR.

Multiplex assays enable the detection of multiple viral RNA and/or DNA targets plus internal controls over a wide linear range without loss of sensitivity (see figures " Reliable detection of viral RNA over a wide linear range" and "   Improved detection of low amounts of viral RNA").

QuantiTect Nucleic Acid Dilution Buffer, supplied with the kits, stabilizes RNA and DNA standards during dilution and reaction setup and prevents loss of nucleic acids on plastic surfaces, such as tubes or pipet tips. The buffer enables reliable dilution of standards used to quantify viral nucleic acids, giving a wide linear range, from low to high CT values and ensures longer storage of standards without degradation (see figure " Reliable dilution and storage of RNA standards").

See figures

Principle

QuantiTect Virus Kits provide highly sensitive detection of viral nucleic acids in single or multiplex assays on the first attempt (see flowchart " QIAGEN multiplex kits"). The optimized master mix ensures that PCR products in a multiplex reaction are amplified with the same efficiency and sensitivity as PCR products in a corresponding single-amplification reaction.

Amplifying control and target genes in the same reaction, instead of separate reactions, increases the reliability of gene quantification by minimizing handling errors. The QuantiTect Virus Buffer contains a balanced combination of K+ and NH4+ ions as well as unique synthetic Factor MP stabilizes, which together promote stable and efficient annealing of primers and probes to the nucleic acid template, enabling high PCR efficiency (see figure " Unique PCR buffer"). In addition, the unique formulation of Sensiscript Reverse Transcriptase ensures highly sensitive reverse transcription of viral RNA, while HotStarTaq Plus DNA Polymerase provides a stringent hot start, preventing the formation of nonspecific products.

Components of the QuantiTect Virus Kit
Kit componentFeature Benefits
5x QuantiTect Virus Master Mix Concentrated master mix Highly concentrated and optimized for sensitive virus detection Larger volumes of template can be added to the assay for increased sensitivity
HotStarTaq Plus DNA Polymerase 5 min activation at 95ºC Set up of qPCR reactions at room temperature
QuantiTect Virus Buffer Balanced combination of NH4+ and K+ ions Specific primer annealing ensures reliable PCR results
Synthetic Factor MP Reliable multiplexing analysis of up to 4 genes in the same tube
Additional kit components QuantiTect Virus RT Mix Contains a unique formulation of Sensiscript Reverse Transcriptase Optimized for highly sensitive detection of viral RNA
QuantiTect Nucleic Acid Dilution Buffer Proprietary buffer formulation for dilution and storage of nucleic acid standards. Stabilizes RNA and DNA standards during dilution and reaction setup and prevents loss of nucleic acids on plastic surfaces, such as tubes or pipet tips
See figures

Procedure

QuantiTect Virus Kits provide highly sensitive real-time PCR analysis of viral nucleic acids (RNA and/or DNA) and internal controls using sequence-specific probes. Reactions can be carried out with or without a reverse-transcription step, enabling flexible design of multiplex assays to detect RNA targets, DNA targets, or both RNA and DNA targets. Follow the protocol in the handbook for fast and reliable results.

Kits are available with or without ROX passive reference dye in the master mix (see table).

Choosing the right QuantiTect Virus Kit
ROX dyeKit Compatible cyclers
Supplied in master mix QuantiTect Virus Kit All cyclers from Applied Biosystems except Applied Biosystems 7500
Supplied in separate tube QuantiTect Virus +ROX Vial Kit Applied Biosystems 7500 and cyclers from
Bio-Rad, Cepheid, Eppendorf, QIAGEN, Roche, Agilent, and other suppliers

For fast and highly sensitive end-point one-step RT-PCR applications, including virus detection, we recommend using the QIAGEN OneStep RT-PCR Kit.

Applications

QuantiTect Virus Kits provide highly sensitive real-time single or multiplex PCR or one-step RT-PCR using sequence-specific probes for detection of viral DNA and/or RNA plus internal controls. The kits can be used on a wide range of real-time thermal cyclers, including instruments from Applied Biosystems, Bio-Rad, Cepheid, Eppendorf, QIAGEN, Roche (except for capillary cyclers), and Agilent.

Supporting data and figures

Specifications

FeaturesSpecifications
applicationsVirus detection
sybrgreeniorsequencespecificprobesSequence-specific probes
realtimeorendpointReal-Time
reactiontypeReverse Transcription and PCR
thermalcyclerMost real-time cyclers (except capillary cyclers e.g. LightCycler® 1.x and 2.0)
sampletargettypeRNA and/or DNA targets
withorwithoutroxAvailable with ROX in master mix and ROX as a separate vial
singleormultiplexSingle or multiplex

FAQ

What is the threshold cycle or Ct value?
The Ct or threshold cycle value is the cycle number at which the fluorescence generated within a reaction crosses the fluorescence threshold, a fluorescent signal significantly above the background fluorescence. At the threshold cycle, a detectable amount of amplicon product has been generated during the early exponential phase of the reaction. The threshold cycle is inversely proportional to the original relative expression level of the gene of interest.
FAQ ID -2682
Why should DNA or cDNA targets be less than 250 bp long for real-time PCR?

Shorter amplification products facilitate high PCR efficiencies. Ideally, amplicon length should be less than 150 bp for optimal amplification efficiency. PCR efficiencies close to 100% are a crucial prerequisite for accurate quantification of target copy numbers in real-time PCR.

FAQ ID-751
What is the difference between QuantiTect Virus Kit and the new QuantiFast Pathogen +IC Kits?
The QuantiTect Virus Kit and the new QuantiFast Pathogen +IC Kits are based on similar reaction chemistry and show identical sensitivity but as a new feature, the QuantiFast Pathogen kits include an Internal Control template and the corresponding primer/probe set for duplex amplification of a user-defined pathogen target with the provided Internal Control. Additional new features are pack size, kit variants, ROX variants, and speed.
FAQ ID -2448
Can the Internal Controls be ordered separately in the QuantiFast Pathogen + IC kit for use during purification?

Yes, the Internal Control RNA (High conc.) and Internal Control DNA (High conc.) templates are available under a separate catalog number. After reconstitution according to the description in the handbook, these IC templates have a 10x higher concentration than the Internal Control templates provided in the kits.   They are sufficiently concentrated to be spiked into the sample prep without replacing too much of the sample input volume.

 

**Please note that there is no assay (primer/probe set) for amplification of the IC included in these separate catalog numbers. The assay is only included in the QuantiFast Pathogen PCR +IC Kit and QuantiFast Pathogen RT-PCR +IC Kit.  The ICs cannot be used with the QuantiTect Virus kits because the assay (primer/probe set) for the detection of the IC is provided with the QuantiFast Pathogen Kits only.

 

FAQ ID -2451