PrimeTime™ qPCR Custom Probe Based Assays
Delivered in convenient single tubes with your specified sequences and optimized primer ratios.
Design your qPCR assays with confidence. Delivered in a single tube or plate well, these assays offer flexible primer-to-probe ratios and use license-free fluorescent dyes. Each component undergoes individual QC—ideal for customizable, high-performance gene expression analysis.
Ordering
- Custom sequence design: Provide your target sequences for probe, forward primer, and reverse primer, or use the PrimerQuest™ Tool to design optimized sequences for any species
- Convenient delivery format: Receive all three oligonucleotides (probe, forward primer, reverse primer) in a single tube or 96-well plate well for streamlined workflow setup
- Flexible primer ratios: Select primer-to-probe ratios from 2:1 to 4:1 with no additional charge to optimize your assay performance
- License-free dye selection: Choose from a wide range of fluorescent dyes including FAM, TET, SUN™, HEX, Cy® 5, Cy® 5.5, Yak™, Texas Red™, ATTO™ 647, and ATTO™ 425
- Multiplex-ready options: Enhance specificity with double-quenched ZEN™ or TAO™ probe assays for confident multiplexing applications
- Digital PCR compatibility: Available in sizes and scales optimized for digital PCR (dPCR) platforms
- Comprehensive documentation: Primer and probe sequence information is provided with every order. Quality control data is available for individual components
PrimeTime qPCR Probe Assays in tubes (1 probe/2 primers)
Available in various sizes, premixed, and shipped dried down.
|
Probes/primers supplied in the following ratios: 0.5/1.0 nmol (Mini); 2.5/2.5–10 nmol (Standard); 12.5/12.5–50 nmol (XL). You may specify the primer-to-probe ratio (except for PrimeTime Mini).
* Predesigned assays are available for human, mouse, and rat targets.
PrimeTime qPCR Probe Assays in 96-well plates (1 probe/2 primers)
Available in various sizes, premixed, and shipped dried down.
|
Probes/primers supplied in the following ratios: 0.5/1.0 nmol (Mini); 2.5/2.5–10 nmol (Standard); 12.5/12.5–50 nmol (XL). You may specify the primer-to-probe ratio (except for PrimeTime Mini). A minimum order of 24 assays is required per plate.
Available dye and quencher combinations for PrimeTime qPCR 5′ Nuclease Assays in plate well format
| 5' dye | 3' quencher | Mini | Standard | XL |
|---|---|---|---|---|
| FAM | ZEN™/Iowa Black™ FQ* | • | • | • |
| FAM | TAMRA | – | • | • |
| HEX | ZEN/Iowa Black FQ* | – | • | • |
| TET | ZEN/Iowa Black FQ* | – | • | • |
| Cy® 5 | Iowa Black RQ | – | • | • |
Key: • = available; – = not available
* ZEN/Iowa Black™ FQ is a Double-Quenched Probe, which provides superior performance compared to traditional single-quenched probes. For more information download the PrimeTime Custom qPCR Probes Flyer.
Request a consultation
Your time is valuable, and we're here to help. If you have questions for our PCR/qPCR/dPCR experts, we’ll prioritize your inquiry to ensure you get the answers you need promptly. Simply click the "Request a Consultation" button, provide some brief information about your project, and our experts will be in touch with you shortly.
Request a consultationProduct details
Our Custom Probe-Based qPCR Assays offer a range of benefits designed to enhance efficiency in your lab. With probes and primers conveniently pre-mixed in a single tube, you can eliminate separate handling steps, saving time and simplifying your workflow. The customizable primer-to-probe ratios allow you to tailor the assay to your specific instrument or experimental needs—whether that’s a 2:1, 3.6:1 or another ratio—without additional cost. This streamlined format also reduces the number of pipetting steps, helping to minimize the risk of contamination and pipetting errors, which provides consistent and reliable results.
Efficient Delivery Format
Our single-tube delivery format streamlines lab workflows by removing the need for separate oligo handling and mixing. This reduces pipetting errors and contamination risks while speeding up assay setup. Customizable primer-to-probe ratios let researchers fine-tune performance for their specific experiments—without extra cost.
Important Considerations
Know Your Ratio
You’ll need to specify your desired primer-to-probe ratio at the time of ordering. If you're unsure, we recommend ordering probes and primers separately so that you can find the ideal ratio for your experimental needs and supports optimization efforts of your new assay design. Explore our PrimeTime qPCR Probes options.
Instrument Compatibility
Some real-time PCR instruments perform best with specific ratios. Be sure to check your instrument’s recommendations before ordering. We also have a PrimeTime™ Multiplex Dye Selection Tool for quick reference.
Universal Features and Research Tools for qPCR Excellence
Tools for Design and Collaboration
This transparency facilitates method sharing and validation across research teams and institutions. For researchers needing help with sequence design, the integrated PrimerQuest™ Tool uses the trusted Primer3 engine with IDT's optimized parameters to generate high-quality designs for endpoint PCR and qPCR across multiple species.
Sequence Information and Manufacturing Expertise
Complete primer and probe sequence information is provided in accordance with MIQE 2.0 recommendations to support reproducible research and publication requirements. IDT's proprietary synthesis platform provides enhanced sequence fidelity and enables incorporation of mixed bases, as well as fluorophores and quencher modifications, resulting in improved quenching efficiency and consistent performance, critical for applications requiring high-confidence detection or comparative analysis across multiple targets.
Customizable, Reliable, and Efficient qPCR Assays
Our Custom Probe-Based qPCR Assays offer flexibility and precision, allowing researchers to tailor their assay to their specific targets. Each assay is rigorously quality-controlled using mass spectrometry* (ESI-MS), with comprehensive QC data available—ensuring transparency and consistent performance. Designed to integrate seamlessly into existing lab workflows, these assays accelerate research by minimizing setup time and preventing signal overlap in multiplex experiments. Their high responsiveness enables identification of gene targets, supporting breakthroughs across diverse fields.
*With the exception of mixed base oligos, which could potentially represent multiple sequences and therefore cannot be evaluated by ESI-MS.
Resources
User guides and protocols
Brochures and flyers
Handbooks
Aids for fluorophore and quencher selection
Gene sets for common pathways
Frequently asked questions
How should I determine the annealing temperature for a probe-based qPCR assay? Should I select a melting temperature midway between my primer and probe melting temperatures (Tm)?
When performing qPCR it is ideal to have your probe Tm about 5−10 degrees higher than your primer Tm. The annealing temperature should be set 3−5 degrees lower than the lowest primer Tm.
Use this as a general guideline, but note that optimization may still be necessary.
What qPCR products does IDT ship at ambient temperature?
We ship PrimeTime™ Gene Expression Master Mix, PrimeTime qPCR assays, and other oligonucleotide products at ambient temperature.
Which dyes are compatible with my thermal cycler?
To determine what dyes are compatible with your instrument, download our Real-time qPCR assay design guide or take a look at the PrimeTime™ Multiplex Dye Selection Tool.
Which dyes are available for use with the IDT ZEN™ quencher?
Currently, the IDT ZEN quencher is available with FAM, HEX, TET, Yakima Yellow® (ELITech Group), JOE™ (Thermo Fisher Scientific), MAX™, and SUN™ fluorophores. Double-quenched probes have significantly lower background fluorescence, especially for longer probes.
The ZEN quencher is available on PrimeTime™ qPCR Probes and on the probes supplied with PrimeTime qPCR Probe Assays. For any special oligonucleotide requests that include the ZEN quencher but are not found on our website (non-catalog requests), please contact us.
For more information, see our DECODED™ article, Double-quenched probes increase qPCR sensitivity and precision.
What tools does IDT provide to design multiplex qPCR assays?
Use the free IDT OligoAnalyzer™ software to help ensure that primer and probe sets lack hairpins and homo- and heterodimer interactions. From there, you can link directly to the NCBI BLAST™ tool to further analyze possible sequence interactions. For help with using the BLAST tool, see Using BLAST to find primers—Tips from IDT.
For more information on setting up multiplex qPCR assays, see the article Recommended dye combinations for multiplex qPCR.
How can I check my PCR primers using the OligoAnalyzer™ program to ensure there are no significant primer design issues?
Look for PCR primers that conform to the following guidelines (use our free online OligoAnalyzer Tool for this purpose):
- The difference between melting temperatures (Tm) of the primers should be less than 5°C.
- The GC content should be between 35−80% or equivalent to the product being amplified.
- The delta G value of any self-dimers, hairpins, and heterodimers should be weaker (more positive) than −9.0 kcal/mol. Positive numbers indicate that the actual secondary structure shown will not form at all.
- Avoid 3' complementarity between the two primers to prevent primer dimers.
The IDT OligoAnalyzer program can be used to assess these different criteria for a proposed oligo.