Agilent Technologies 4150 TapeStation system User Manual

Agilent 4150 TapeStation System Manual 201
Troubleshooting
10
Troubleshooting RNA Applications
Sample not collected
at bottom of vial
Insufficient volume pipetted by the robotics and
applied to the lane may affect the migration
behavior and alter the band shape.
Make sure to collect all liquid at the bottom of
the vial by centrifugation. See “Centrifugation
Recommendations” on page 97. Ensure that
after removing strip caps and inserting the tube
strips or 96-well sample plate into the
TapeStation instrument liquid is still located at
the bottom of the well, and is not spilled.
Incorrect peak
integration
The algorithm cannot establish a correct
concentration estimation if marker or sample
peaks are not completely encompassed
because the signal from either marker or
sample is left out of the calculation.
Ensure that all sample peaks and markers are
integrated correctly in the TapeStation Analysis
software, by clicking and dragging so that the
whole peak is encompassed, see “Peak
Integration” on page 101.
See the Agilent Information Center for more
information.
Incorrect Marker
peaks picked up
Depending on the sample pattern wrong peaks
can get assigned as marker if running very
close to the actual marker.
Use the Aligned button to detect the correct
markers. Compare to aligned/unaligned data of
good, for example the ladder, and questionable
samples, then assign correct markers by
right-clicking.
See the Agilent Information Center for more
information.
Ensure that all sample peaks are within the
recommended “Sizing Range” on page 106 for
the application.
Incorrect heating
procedure
Both over and under-heating the samples can
affect concentration values due to differences
in the migration behavior of RNA still trapped in
secondary structure.
Ensure that the samples are heat denatured
according to the assay instructions before
running.
Sample and the
marker relevant for
quantification have
merged.
Additional sample overlaying with the marker
will alter the known concentration of the Lower
Marker.
For these samples an accurate quantification
cannot be provided. Highly concentrated
samples may be diluted in order to minimize the
amount of sample merging with the Lower
Marker.
Contaminating beads can alter the run behavior
of the sample and can introduce artifacts.
If using the Sure Select Protocol, ensure that all
AMPure beads are removed by increasing the
time that the sample is on the magnetic plate.
Table 11 RNA - Possible causes of incorrect quantification in order of probability.
Root cause Explanation Solution

Agilent Technologies 4150 TapeStation system Specifications

General

Brand	Agilent Technologies
Model	4150 TapeStation system
Category	Laboratory Equipment
Language	English