public void TestSinglePointExternalCalibrationWithIsConcentration()
{
const double internalStandardConcentration = 80.0;
var srmDocument = LoadTestDocument();
srmDocument = ChangeStandardConcentrationCount(srmDocument, 1);
srmDocument = ChangeQuantificationSettings(srmDocument, QuantificationSettings.DEFAULT
.ChangeRegressionFit(RegressionFit.LINEAR_THROUGH_ZERO)
.ChangeNormalizationMethod(NormalizationMethod.GetNormalizationMethod(IsotopeLabelType.heavy)));
PeptideGroupDocNode protein = srmDocument.MoleculeGroups.First();
PeptideDocNode peptide = protein.Peptides.First();
peptide = peptide.ChangeInternalStandardConcentration(internalStandardConcentration);
PeptideQuantifier peptideQuantifier = PeptideQuantifier.GetPeptideQuantifier(srmDocument.Settings, protein, peptide);
CalibrationCurveFitter curveFitter = new CalibrationCurveFitter(peptideQuantifier, srmDocument.Settings);
Assert.AreEqual(CalibrationCurveFitter.ConcentrationRatioText(IsotopeLabelType.light, IsotopeLabelType.heavy),
curveFitter.GetXAxisTitle());
Assert.AreEqual(CalibrationCurveFitter.PeakAreaRatioText(IsotopeLabelType.light, IsotopeLabelType.heavy), curveFitter.GetYAxisTitle());
var chromatograms = srmDocument.Settings.MeasuredResults.Chromatograms;
for (int iReplicate = 0; iReplicate < chromatograms.Count; iReplicate++)
{
string msg = string.Format("Replicate {0}", iReplicate);
var expectedY = peptide.Results[iReplicate].First().LabelRatios.First().Ratio.Ratio;
double?actualY = curveFitter.GetYValue(iReplicate).Value;
Assert.IsNotNull(actualY);
Assert.AreEqual(expectedY, actualY.Value, epsilon, msg);
var calibrationCurve = curveFitter.GetCalibrationCurve();
Assert.AreEqual(1, calibrationCurve.PointCount);
var expectedConcentration = expectedY * internalStandardConcentration / calibrationCurve.Slope.Value;
Assert.AreEqual(expectedConcentration / internalStandardConcentration,
curveFitter.GetCalculatedXValue(calibrationCurve, iReplicate).GetValueOrDefault(double.NaN), epsilon, msg);
Assert.AreEqual(expectedConcentration,
curveFitter.GetCalculatedConcentration(calibrationCurve, iReplicate).GetValueOrDefault(double.NaN), epsilon, msg);
var chromatogramSet = chromatograms[iReplicate];
if (Equals(chromatogramSet.SampleType, SampleType.STANDARD) && chromatogramSet.AnalyteConcentration.HasValue)
{
// Since there was only one standard, the regression line goes exactly through this sample.
Assert.AreEqual(expectedConcentration, chromatogramSet.AnalyteConcentration.Value * peptide.ConcentrationMultiplier.GetValueOrDefault(1.0), epsilon);
}
}
}
public void TestInternalCalibration()
{
const double internalStandardConcentration = 80.0;
var srmDocument = LoadTestDocument();
AssertEx.IsDocumentState(srmDocument, null, 1, 1, 2, 10);
srmDocument = ChangeStandardConcentrationCount(srmDocument, 0);
srmDocument = ChangeQuantificationSettings(srmDocument,
QuantificationSettings.DEFAULT
.ChangeRegressionFit(RegressionFit.NONE)
.ChangeNormalizationMethod(NormalizationMethod.GetNormalizationMethod(IsotopeLabelType.heavy))
.ChangeUnits("ng/mL"));
PeptideGroupDocNode protein = srmDocument.MoleculeGroups.First();
PeptideDocNode peptide = protein.Peptides.First();
peptide = peptide.ChangeInternalStandardConcentration(internalStandardConcentration);
CalibrationCurveFitter curveFitter = CalibrationCurveFitter.GetCalibrationCurveFitter(srmDocument.Settings, protein, peptide);
string expectedXAxis = CalibrationCurveFitter.AppendUnits(QuantificationStrings.Analyte_Concentration, "ng/mL");
Assert.AreEqual(expectedXAxis, curveFitter.GetXAxisTitle());
string expectedYAxis = CalibrationCurveFitter.PeakAreaRatioText(IsotopeLabelType.light, IsotopeLabelType.heavy);
Assert.AreEqual(expectedYAxis, curveFitter.GetYAxisTitle());
var chromatograms = srmDocument.Settings.MeasuredResults.Chromatograms;
for (int iReplicate = 0; iReplicate < chromatograms.Count; iReplicate++)
{
string msg = string.Format("Replicate {0}", iReplicate);
var expectedY = peptide.Results[iReplicate].First().LabelRatios.First().Ratio.Ratio;
Assert.AreEqual(expectedY, curveFitter.GetYValue(iReplicate).Value, epsilon, msg);
var calibrationCurve = curveFitter.GetCalibrationCurve();
Assert.AreEqual(1 / internalStandardConcentration, calibrationCurve.Slope);
Assert.IsNull(calibrationCurve.RSquared);
Assert.AreEqual(expectedY * internalStandardConcentration,
curveFitter.GetCalculatedXValue(calibrationCurve, iReplicate).GetValueOrDefault(double.NaN), epsilon, msg);
Assert.AreEqual(expectedY * internalStandardConcentration,
curveFitter.GetCalculatedConcentration(calibrationCurve, iReplicate).GetValueOrDefault(double.NaN), epsilon, msg);
}
}
protected override void DoTest()
{
RunUI(() => SkylineWindow.OpenFile(TestFilesDir.GetTestPath("p180test_calibration_DukeApril2016.sky")));
var surrogateStandards = new Dictionary <string, string>
{
{ "Leu", "Ile" },
{ "Lys", "Orn" },
{ "Ac-Orn", "Orn" },
{ "SDMA", "ADMA" },
{ "alpha-AAA", "Orn" },
{ "Carnosine", "His" },
{ "Histamine", "His" },
{ "Kynurenine", "Tyr" },
{ "Met-SO", "Met" },
{ "Nitro-Tyr", "Tyr" },
{ "c4-OH-Pro", "Pro" },
{ "t4-OH-Pro", "Pro" },
};
// Set the standard type of the surrogate standards to StandardType.SURROGATE_STANDARD
RunUI(() =>
{
List <IdentityPath> pathsToSelect = SkylineWindow.SequenceTree.Nodes.OfType <PeptideGroupTreeNode>()
.SelectMany(peptideGroup => peptideGroup.Nodes.OfType <PeptideTreeNode>())
.Where(peptideTreeNode => surrogateStandards.Values.Contains(peptideTreeNode.DocNode.RawTextId))
.Select(treeNode => treeNode.Path)
.ToList();
SkylineWindow.SequenceTree.SelectedPaths = pathsToSelect;
SkylineWindow.SetStandardType(StandardType.SURROGATE_STANDARD);
});
// Use the document grid to set the Normalization Method of the molecules that have surrogate standards
var documentGrid = ShowDialog <DocumentGridForm>(() => SkylineWindow.ShowDocumentGrid(true));
RunUI(() => documentGrid.ChooseView(Resources.Resources_ReportSpecList_GetDefaults_Peptide_Quantification));
IDataGridViewEditingControl editingControl = null;
DataGridViewEditingControlShowingEventHandler onEditingControlShowing =
(sender, args) => editingControl = args.Control as IDataGridViewEditingControl;
documentGrid.DataGridView.EditingControlShowing += onEditingControlShowing;
// ReSharper disable AccessToForEachVariableInClosure
foreach (var entry in surrogateStandards)
{
WaitForConditionUI(() => documentGrid.IsComplete);
RunUI(() =>
{
var colPeptide = documentGrid.FindColumn(PropertyPath.Root);
for (int iRow = 0; iRow < documentGrid.RowCount; iRow++)
{
var row = documentGrid.DataGridView.Rows[iRow];
if (!entry.Key.Equals(row.Cells[colPeptide.Index].FormattedValue))
{
continue;
}
var colNormalizationMethod = documentGrid.FindColumn(PropertyPath.Root.Property("NormalizationMethod"));
documentGrid.DataGridView.CurrentCell = row.Cells[colNormalizationMethod.Index];
documentGrid.DataGridView.BeginEdit(false);
editingControl.EditingControlFormattedValue =
string.Format(Resources.RatioToSurrogate_ToString_Ratio_to_surrogate__0____1__, entry.Value,
"Heavy");
documentGrid.DataGridView.EndEdit();
}
});
}
// Make sure that the Y-Axis on the CalibrationForm reflects the normalization method of the selected molecule
var calibrationForm = ShowDialog <CalibrationForm>(() => SkylineWindow.ShowCalibrationForm());
foreach (var peptideGroupTreeNode in SkylineWindow.SequenceTree.Nodes.OfType <PeptideGroupTreeNode>())
{
foreach (var peptideTreeNode in peptideGroupTreeNode.Nodes.OfType <PeptideTreeNode>())
{
RunUI(() => SkylineWindow.SequenceTree.SelectedPath = peptideTreeNode.Path);
WaitForGraphs();
string yAxisText = calibrationForm.ZedGraphControl.GraphPane.YAxis.Title.Text;
if (null != peptideTreeNode.DocNode.NormalizationMethod)
{
Assert.IsInstanceOfType(peptideTreeNode.DocNode.NormalizationMethod, typeof(NormalizationMethod.RatioToSurrogate));
Assert.AreEqual(QuantificationStrings.CalibrationCurveFitter_GetYAxisTitle_Normalized_Peak_Area,
yAxisText);
}
else
{
Assert.IsInstanceOfType(SkylineWindow.Document.Settings.PeptideSettings.Quantification.NormalizationMethod, typeof(NormalizationMethod.RatioToLabel));
string expectedTitle = CalibrationCurveFitter.PeakAreaRatioText(IsotopeLabelType.light,
IsotopeLabelType.heavy);
Assert.AreEqual(expectedTitle, yAxisText);
}
}
}
}