• Freeze-dryable formulation including excipients and stabilizers
• Best-in-class performance in both single and multiplex quantitative amplification
• Convenient master mix for low-copy-number pathogen detection
• High specificity and sensitivity across a wide range of sample sources
• Minimized risk of carryover contamination
• Compatible with a wide range of probe technologies, including TaqMan®, Molecular Beacons® and Scorpion® probes

• PCR amplification of viral DNA, cDNA and genomic DNA targets
• For use with standard and fast qPCR platforms
• Single and multiplex qPCR amplification
• Suitable for high-throughput laboratories

biotechrabbit™ 4X LYO-ready CAPITAL™ qPCR Probe Master Mix Plus is a ready-to-use, four-times-concentrated formulation containing an adjusted concentration of heat-labile Uracil-DNA Glycosylase (HL-UDG) to minimize the risk of carryover contamination. The mix includes all enzymes, dNTPs, excipients and stabilizers required for producing in-house lyophilizates, with the option to include primers and probes. Time-efficient lyophilization is possible for both high and low volumes of the master mix without collapse of the lyophilizate. The resulting lyophilizate is highly resistant to humidity and dissolves rapidly in the provided reconstitution buffer.

The master mix is optimized for real-time PCR quantification of viral DNA, cDNA and genomic DNA from a wide range of targets. It ensures high specificity and sensitivity in single and multiplex amplification, making it an excellent choice for extremely low-copy-number targets in pathogen detection.

4X LYO-ready CAPITAL™ qPCR Probe Master Mix Plus uses a proprietary enzyme and buffer formulation suitable for fast extension and multiplexing in challenging PCR reactions.

Handling

Prevention of contamination

Contamination with undesired DNA is a concern when assembling amplification reactions. To minimize the risk of contamination, follow the guidelines below:

• Wear disposable gloves when handling the solutions.
• Dedicate separate sterile areas for the preparation of samples and reaction mixtures.
• Use molecular-grade nuclease-free water and reagents.
• Include a non-template control reaction in every PCR assay.

Recommendations before starting a lyophilization project

• The formulation is optimized for freeze-drying both single and multiple reactions with a reaction volume of 20 µl.
• Optional: Prepare concentrated primers and probes in molecular-biology-grade water and restrict their volume to a maximum of 20 % of the LYO-ready master mix.

Calculation examples

Three calculation examples for a 20 µl reaction size:

Pipetting scheme

Divide the total reaction volume above by a factor of 4. Minimize the volume of primers and probes. Do not add water.

*Depending on the pipetting volume.

Suggested lyophilization protocol

*The duration of freeze-drying cycles depends on the lyophilizate volume, the equipment, and the type and dimensions of the vials.

Reconstitution of lyophilized master mix

• Add the required volume of reconstitution buffer to the lyophilized CAPITAL™ qPCR Probe Master Mix Plus, as shown in the examples below.
• Mix well. The lyophilizate should dissolve rapidly.
• Store the reconstituted CAPITAL™ qPCR Probe Master Mix Plus at −20 °C.

Cycling program

*The recommended annealing/extension temperature is primer Tm +2 °C. Use gradient PCR to optimize the annealing temperature. Do not use annealing temperatures below 60 °C.