AUGUST | SEPTEMBER 2013 Forensic Magazine | wwww.forensicmag.com 15
tion level more closely to the DNA profile generated
with NG STR PCR amplification kits. The collective
data indicates that this new degradation method, when
assessed along with the IPC results, provides forensic
laboratories a DNA QI that better informs their decisions on how to optimize STR PCR setup to enhance
efficiency and increase first pass success rates of STR–
based DNA profiling for challenging samples.
As a result of technology advancements, including
the optimized PCR buffer composition and selected
human and male targets, an increased correlation is
obtained between DNA quantities and downstream
DNA profiles using NG STR PCR amplification kits.
Especially, high sensitivity and accuracy at lower DNA
concentrations (5–100 pg/µl range) is important to obtain reliable DNA quantification data for low template
DNA samples. In order to reliably generate STR–based
DNA profiles with a low probability of allelic dropout,
forensic laboratories often set a DNA quantity threshold at the lower DNA concentration range to allow
for maximum input of DNA within the PCR reaction.
Therefore, accurate DNA quantification data is of great
importance to meet enhanced efficiency and first pass
success rates. During developmental validation studies and verification testing, accurate determination of
the amount of DNA was obtained for a DNA dilution
series ranging from more than 100 ng/µl to less than 5
pg/µl. Although stochastic noise tends to compromise
the precision of quantification measurements below 5
pg/µL, the improved sensitivity of the kits allows DNA
concentrations even at the sub-picogram range to be
detected. Most importantly, testing of various casework
samples showed an improved correlation between DNA
quantities below 100 pg/µl and the resultant DNA
profile. DNA samples previously showing “0” (
undetermined) DNA quantity values with other quantification
kits, but showing DNA profiles upon NG STR PCR
amplification, show a concordant DNA quantification
result with NG quantification kits.
High sensitivity for the Y target is of major importance to detect male contribution in traces related to
sexual assault, where usually mixed traces contain DNA
from both the male perpetrator and (often in excess) of
the female victim. Mixture analysis studies performed
during developmental validation have shown a high
sensitivity of detection of male contribution in the presence of more than a 1000-fold excess of female DNA.
Further analysis between PCR inhibitors and ef-
ficiency of PCR amplification of the IPC target reveal
that higher inhibitor tolerance is obtained and mirrors
results to next generation STR PCR amplification kits.
Finally, during development validation studies, the
tolerance of quantification kits was investigated. The
Human Acid and ten times higher tolerance for Hema-
tin similar to the tolerance obtained by with the latest
NG STR PCR amplification kits.
Collected data demonstrates that the NG quantification kits provide valuable sample information on the
quantity and quality of DNA, and are helpful in informing critical decisions that increase the success rate of
obtaining interpretable DNA profiles in subsequent
analysis. With the significant reduced time required to
set up the assay and the enhanced performance, these
quantification kits can be successfully implemented as a
fast decision making tool to obtain more DNA related
information for criminal investigations more quickly.
Next Generation Quantification Kits Streamline
Decision Making Process
Over the last year, the development of more robust
DNA quantification and assessment kits provides better
correlation between the DNA sample quantity and quality with the resultant STR profile. These next generation DNA quantification and assessment kits have high
sensitivity (sub-pg level), for both the human and male
targets, higher inhibitor tolerance to match next generation STR kits, and additional critical tools for the determination of DNA quality. In addition, the time required
to set up the assay and perform amplification has been
significantly reduced compared to previous quantification
methods. Further, the standard curve generation protocol
is optimized to provide consistent results. This system is
a decision making tool and has correlative abilities that
have been successful in obtaining STR profiles from challenging samples.
3.Barbisin et al., Journal of Forensic Sciences, 54, 305–319
Wiljo De Leeuw joined Life Technologies in 2009 as
Senior Field Application Specialist for EMEA Nordic in the
HID Applied Markets.
Sheri Olson is a Global Sr. Product Manager for the Human Identification business at Life Technologies.
Robert L. Green joined the Human Identification Group
at Life Technologies in 1996, where he developed the first
generation of Quantifiler Kits.
Allison Holt is an R&D Scientist in the Human Identification business at Life Technologies. email@example.com
Lisa Lane Schade is the Director of Global Market
Development for the Human Identification Business at Life