Calibrate Custom Dyes; Custom Dyes Overview; Use A Dilution Series To Determine An Optimal Custom Dye Concentration - Thermo Scientific QuantStudio 1 Installation, Use And Maintenance Manual

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Chapter 5 Calibrate and verify instrument performance

Calibrate custom dyes

Calibrate custom dyes

Custom dyes
overview
Use a dilution
series to
determine an
optimal custom
dye concentration
62
The instrument can run assays designed with custom dyes. Custom dyes include:
• Dyes that are not manufactured by Thermo Fisher Scientific.
• Dyes or formulations of dyes that are not system dyes for the instrument.
• For custom dye requirements, see "Custom dyes" on page 10.
Workflow: Custom dye calibration
For each custom dye, determine the optimal dye concentration. Use this concentration
to prepare all subsequent dye calibration plates.

Use a dilution series to determine an optimal custom dye concentration

"Prepare a custom dye dilution plate" on page 63
"Run the dilution plate as an experiment" on page 64
"Determine the optimal dye concentration" on page 64
Calibrate the custom dye using the optimal concentration
"Perform a custom dye calibration" on page 66
Custom dye dilution guidelines
Prepare a dilution series for each custom dye.
• Target several dye concentrations within a range of 100–2,000 nM.
• Choose a 2- or 3-fold difference in dilution points.
• Dispense 10–20 µL per well in a 96-well plate.
• Dilute the dye in buffer compatible with your master mix.
• (Intercalating dyes only) Add the appropriate amount of amplified PCR product to
generate fluorescence.
QuantStudio
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Create a custom dye calibration plate
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Add a custom dye to the instrument
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1 Real-Time PCR System Installation, Use, and Maintenance Guide
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