LabChip GX/GXII User Manual

Glossary of Terms 261Polynomial OrderThis setting on the Peak Find Tab determines the order of thepolynomial filter used to convolve with the data. A polynomial filteris used to filter the data to increase the signal-to-noise ratio andcalculate filter slope information for peak detection. The defaultsetting is 6 (for 6th order). A setting of 4 produces a straighterbaseline, preventing quick changes in the signal.Pressurization SyringeA plastic syringe, which is attached to the chip priming station. Thissyringe can be used to force the buffer solution into all the channelsin the chip prior to running the chip in the instrument.Protein Assay AnalysisProtein assays utilize a chemistry that generates an extra set ofpeaks (system peaks) just above the lower marker that should notbe included in the analysis. These system peaks must be identifiedand excluded from further quantitative analysis. For ladder analysis,peaks occurring after the lower marker less than the “Ladder Ratio”analysis setting, are tagged system peaks to avoid confusion withlegitimate ladder peaks.Protein samples contain a single lower marker. Alignment to asingle marker does not provide enough constraints to align largeproteins, so data is aligned to two ladders, one sipped just beforethe sample wells and another sipped just after the sample wells.The second ladder is called the bracket ladder. Samples are scaledso the sample’s lower marker is nearly aligned with both ladderlower markers. The scaling is weighted by the proximity in sip timeto each ladder. The sample sipped closest to the primary ladder isscaled to align more closely to the primary ladder and the samplesipped just before the bracket ladder is scaled to align most closelyto the bracket ladder. Then each sample is shifted in time so thesample's lower marker aligns exactly with the primary ladder lowermarker.After alignment, the size of the protein producing each peak iscalculated from the aligned peak time using a log (size) versus1/(Time-T0) fit to the primarily ladder peaks of known size andmeasured migration time. T0 is determined empirically as the timeoffset which delivers the best straight line fit to the ladder data, Thevalue of T0 used for the fit can be viewed in the Well Table View.P/N 127170 Rev. 00 LabChip GX User Manual Caliper Life Sciences, Inc.