myT BRAF-Ultra

Product Name Catalog No. Price*
myT BRAF-Ultra BRAF-U1 $1,495.00 USD
qPCR primers for ultra-low copy number detection of BRAF mutations V600E/K

Provides superior sensitivity and high confidence results:

  • 0.01% sensitivity with very low background amplification
  • Single copy detection capability
  • Ideal for circulating tumor cells (CTCs), serum, plasma and needle biopsies



Each package includes reagents to perform a total of 60 reactions (16 locus-specific reactions and 44 allele-specific reactions performed in triplicate), sufficient for testing up to 14 samples plus a positive and no template control per primer cocktail if performed as a single qPCR run.

The detection probe and PCR master mix must be ordered separately from licensed vendors. The purchase price of the probe and shipping costs are included in your purchase of myT BRAF-Ultra. Instructions on how to easily order the probe from our recommended vendor and a coupon covering the purchase price and shipping cost of the probe will be provided with your order acknowledgement. This will enable you to synchronize delivery of the probe with delivery of myT BRAF-Ultra. The cost of master mix purchase is not included. Please refer to myT BRAF-Ultra Product Details for recommended vendors and additional information.

Product Name Catalog No. Price*
myT BRAF-Ultra BRAF-U1 $1,495.00 USD

*Inquire about quantity discounts.


Notice to Purchaser: Limited License
This product is for research use only and is licensed to the user under Swift Biosciences intellectual property only for the purchaser’s internal research. Purchase of this product does not convey to the purchaser any license to perform the Polymerase Chain Reaction “PCR” process under any third party rights. PCR probes can be purchased from a variety of vendors including Applied Biosystems (Life Technologies), Roche Molecular Systems, Inc., F. Hoffman La-Roche Ltd., Integrated DNA Technologies, Biosearch Technologies, Nanogen Inc. and others. The use of certain probes including TaqMan-MGB, FAM-TAMRA, FAM-BHQ, VIC-MGB in connection with ("PCR") process may require a license from one or more of these vendors. Please contact individual vendors for the necessity of obtaining licenses. The purchase of myT BRAF or any other items delivered by Swift Biosciences hereunder does not, either expressly or by implication, provide a license to use any proprietary technology of these vendors.


Mutation detection with myT BRAF-Ultra consists of two steps:
  1. Locus-specific qPCR
    A non-allele-specific qPCR is performed to assess total (mutant + wild-type) amplifiable BRAF for each sample. This determines the quantity of DNA to be used in step 2 for each sample.

  2. Allele-specific qPCR
    A mutant allele-specific BRAF qPCR is then performed to assess presence of V600E or V600K. This assay DOES distinguish between wild-type and mutant but DOES NOT distinguish between the two mutant alleles. Results are reported as positive or negative for mutant BRAF for each sample with a sensitivity limit of 0.01%. This corresponds to one mutant copy in the presence of 10,000 wild-type copies.

    Note: Due to the unavoidable Poisson distribution of ultra-low templates (down to single copy), it is possible that any single reaction well may have zero, one or more templates. As a result, triplicate analysis is required to ensure detection when the template copy number per reaction is expected to be low (<10 copies per reaction).

myT BRAF-Ultra Workflow

Isolate genomic DNA from samples


Obtain UV absorbance readings

 

Perform locus-specific qPCR

 

Determine amplifiable copy number from Ct values

 

Perform allele-specific qPCR

 

Determine BRAF V600E/K status

Reagents Included
Reagent Mix Description
BRAF-Ultra Locus-specific Primers  Non-allele-specific primers
BRAF-Ultra Allele-specific Primers
V600E/K allele-specific myT BRAF-Ultra primers
Nuclease-free Buffer For DNA sample dilution and NTC reactions
BRAF DNA Standard BRAF V600E control DNA (200 copies/µl)


Reagents Not Included
The following reagents are not provided and must be ordered separately from a licensed vendor. The purchase price of the probe and shipping costs are included in your purchase of myT BRAF-Ultra. Instructions on how to easily order the probe from our recommended vendor and a coupon covering the purchase price and shipping cost of the probe will be provided with your order acknowledgement. This will enable you to synchronize delivery of the probe with delivery of myT BRAF-Ultra. The cost of master mix purchase is not included.

Reagent Recommended Vendor
qPCR Master Mix (2X) Fermentas Maxima Probe qPCR Master Mix, Catalog # K0261
Detection Probe IDT PrimeTime Dual-Labeled Probe (voucher included with purchase)

Number of Reactions
The product contents are sufficient to perform 60 reactions consisting of 16 locus-specific single reactions and 44 allele-specific reactions (samples run in triplicate). This enables testing of up to 14 samples when including a positive control and a no-template control (NTC) when performed as a single qPCR run. If testing is split into multiple batches, total samples tested will be less as a positive control and NTC are recommended for each run.

Product Literature
Material Safety Data Sheets
PDF versions of Material Safety Data Sheets (MSDS) are available for download.

myT BRAF-Ultra Performance on the ABI 7500

Top: qPCR reactions containing mutant DNA at the specified quantity in a background of 14,000 copies of wild-type genomic DNA resulted in BRAF V600E-specific amplification: 1% mutant content (purple; n = 8 replicates), 0.1% mutant content (green; n = 8 replicates) and 0.01% mutant content (blue; n = 8 positive replicates).

Bottom: qPCR reactions containing 14,000 copies of wild-type genomic DNA (no mutant DNA) resulted in 2% non-specific amplification (n = 95 replicates).




Conclusion: These results demonstrate mismatch discrimination with 100% sensitivity and 98% specificity. Wild-type DNA demonstrates very low non-specific amplification (2%) and a single mutant copy is reliably detected 100% of the time (taking Poisson distribution into account). This capability in ultra-low copy number detection is a feature that is exclusive to myT Primer-Ultra reagents.

*For research use only

Trademarks
ABI 7500 is a product of Applied Biosystems, now part of Life Technologies Corporation