User manual

28 | Droplet Digital
™
PCR Applications Guide
3 Absolute Quantification
and the Statistics of
Droplet Digital
™
PCR
Running Absolute Quantification Experiments
Absolute quantification (ABS) is fundamental to all Droplet Digital PCR (ddPCR
™
)
applications. Partitioning template DNA into uniform droplets enables highly quantified
measurements of target DNA using the QX200
™
or QX100
™
Droplet Digital PCR system
and the appropriate automatic statistical analysis in QuantaSoft
™
software. ddPCR does
not require a standard curve, and this simplifies the experimental planning and procedures,
and reduces result variability and cost.
Important ABS experiment considerations are as follows:
■
Digest the template DNA to fragment the genome. Select restriction enzymes that
do not cut either the target or reference amplicons
■
The restriction-digested DNA should be at a higher concentration in order to have
a lower concentration in the master mix
Note: Account for the concentration of the template in any restriction digest and the
amount of template in the final 1x ddPCR supermix to backcalculate the original
concentration of the stock template.