Search Thermo Fisher Scientific
Search Thermo Fisher Scientific
Catalog Number | No. of Reactions |
---|---|
A25603 | 100 Reactions |
A25602 | 400 Reactions |
A25605 | 20 Reactions |
Like other members of the Cells-to-CT™ product family, the Cells-to-CT™ 1-Step TaqMan™ Kit makes it possible to perform expression analysis directly from cultured cells without RNA purification.
This new addition to the family includes a TaqMan™ 1-Step RT-PCR master mix. The single-tube format of the RT-PCR step streamlines the 'sample to answer' workflow more than ever. Combining the simple Cells-to- CT™ lysis technology with the fast cycling capability of the 1-step RT-PCR master mix allows quicker time to results with limited sample handling.
The Cells-to-CT™ 1-Step TaqMan™ Kit is:
Note: TaqMan™ primer/probe sets are sold separately.
Simple 7-minute Sample Preparation, Then Ready for Real-time RT-PCR
Whether you are using plates or tubes, the Cells-to-CT™ 1-Step TaqMan™ Kit uses a simple 7-minute sample preparation procedure. The lysis technology is designed for 10–100,000 cultured cells/sample. Cells are washed in PBS and lysed in solution for 5 minutes at room temperature with a simultaneous DNase treatment. Lysis is terminated at room temperature by a 2-minute incubation with Stop Solution. The lysate is now ready to analyze on your real-time PCR instrument. Simply add it to a real-time RT-PCR master mix containing the TaqMan™ primer/probe sets of interest. Alternatively, the lysate can be stored at –20°C. Because samples can be processed directly in culture wells (96 or 384 wells), sample handling and the potential for sample loss or transfer error are minimized, facilitating rapid, high-throughput processing. Unlike old-fashioned, multi-step RNA isolation protocols, no heating, washing, or centrifugation steps are required. The kit greatly simplifies a laborious 30–60 minute process and reduces it to 7 minutes.
A Complete, Validated Solution
All components of the Cells-to-CT™ 1-Step TaqMan™ Kit have been optimized for consistent and reliable performance. This removes the guesswork involved in assembling separate sample preparation and real-time 1-step RT-PCR master mixes. For added quality assurance, the Cells-to-CT™ 1-Step TaqMan™ Kit has been validated with TaqMan™ Gene Expression assays and shows performance equivalent to that obtained with purified RNA.
Accessory Product
The TaqMan™ Cells-to-CT™ Control Kit was designed for use with TaqMan™ Cells-to-CT™ products. This kit contains an endogenous control (ACTB) to normalize for sample loading variability and an artificial XenoRNA™ control and corresponding TaqMan™ Gene Expression assay to monitor the effects of PCR inhibition.
If you are targeting a low-abundance gene, you may have trouble getting Ct values in a good, reliable range (Ct > 32). To increase the sensitivity of the assay, you may want to consider the following:
- Increase the amount of RNA input into your reverse transcription reaction, if possible
- Increase the amount of cDNA in your qPCR reaction (20% by volume max)
- Try a different reverse transcription kit, such as our SuperScript VILO Master Mix, for the highest cDNA yield possible
- Consider trying a one-step or Cells-to-CT type workflow (depending on your sample type)
Most times your instrument software can automatically set a proper baseline for your data. Check out our short video, Understanding Baselines, for more information on how to set them (https://www.youtube.com/watch?feature=player_embedded&v=5BjFAJHW-bE).
In most cases your instrument software can automatically set a proper threshold for your data. Check out our short video, Understanding Thresholds, for more information on how to set them (https://www.youtube.com/watch?feature=player_embedded&v=H_xsuRQIM9M).
There could be several reasons for no amplification from an assay or primer set. Please see these examples and suggested solutions in our Real-Time Troubleshooting Tool (https://www.thermofisher.com/us/en/home/life-science/pcr/real-time-pcr/qpcr-education/real-time-pcr-troubleshooting-tool/gene-expression-quantitation-troubleshooting/no-amplification.html) for more details.
There could be several reasons for amplification in a NTC well. Please see these examples and suggested solutions in our Real-Time Troubleshooting Tool (https://www.thermofisher.com/us/en/home/life-science/pcr/real-time-pcr/qpcr-education/real-time-pcr-troubleshooting-tool/gene-expression-quantitation-troubleshooting/amplification-no-template-control.html) for more details.
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