Clarity User Training

1
Clarity User Training

• Basic operations

2
Basic Operations

• Method development
• Integration parameters
• Calibration
• Reports

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Opening the station
Status table
Instrument window
Analysis(Single run/Sequence)
Method Setup
Clarity Main window
Data Acquisition
Chromatogram
Report
Calibration
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Template method concept

• Template Method opened on Instrument serves as a
  template for new chromatograms
• The content is copied to chromatogram after
  acquisition or batch reprocessing
• Chromatogram method
• Changes in chromatogram methoddo not affect the
  template method
• Linked Calibration
• Calibration file is linked to method (template
  or chromatogram) by its name

Template Method opened on Instrument
Template Method used for current run
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Clarity Method development

• Template Method parameters
• Sample information
• Acquisition of a typical chromatogram (standard)
• Integration parameters setting
• Creating a calibration file
• Acquisition of other standards and unknown
  samples
• Printing and export of results

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Measurement conditions

• Measuring conditions
• informative, quite useful after time (when filled
  in)
• Autostop setting
• necessary for Active Sequence

External start signal options
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Acquisition

• Data acquisition parameters
• Range, Sampling Rate individual for each
  detector
• Range and Sample Rate
• critical settings, if wrong, analysis must be
  repeated

Select/enable signal (detector)
Caution this parameters cannot be changed later!
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Range and Sample Rate

• Range
• according to detector output
• signal outside range is lost, wrong areas and
  heights
• signal sometimes limited by detector
• Sample Rate
• for capillary GC set 25-50 Hz, otherwise 10 Hz
• when low, insufficient data points for peak
  description

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Method Setup - Integration
Editable Integration table with default parameters
Individual table for each signal
To set up this table properly measured
chromatogram is necessary. Easiest way is to use
the graphical tools in Chromatogram window
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Method - Calculation
Set name of existing calibration file(or create
new file)
The Calibration file is linked to method and
chromatograms by its name. It is advisable to set
it in the method even before any chromatogram is
measured.
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Sample info
Sample description and Sample ID informative
fields
Values used in calibrated calculations
Snapshot process the data acquired so far
without finishing the running analysis
Store file in CALIB or DATA subdirectory Recalibra
te selected level
File Name variables for automatic name composing
Existing files will be overwritten without
warning
RAW data processing from TMP subdirectory
Initial value for n
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Sequence table window
File name, automatic variables for SV, EV, IV
Automatic recalibration from sequence
Individual Post-Run settings for each row
Start, End vial and Injections number. Several
vials and/or injections defined on single row
Tooltip with actual file name
Use row, add new rows during sequence run
Row status indicator
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Chromatogram acquisition

• Inject sample
• Start the data acquisition
• automatically, by external start cable connected
  to valve
• manually, by button on instrument or start cable
• manually, the RUN button in Single Analysis
  dialogor icon in Data Acquisition window
• Watch the acquired signal in the Data acquisition
  window
• Stop the data acquisition
• automatically, after the Autostop time set
• manually, the Stop button in Single Analysis
  dialogor icon in Data Acquisition window

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Data Acquisition
Baseline monitoring (before analysis)
Analysis monitoring
Display only, not influencing collected data
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Integration Table Parameters

• Optimize Peak Width and Threshold
• basic parameters for integration algorithm
• Baseline and peak modifications
• integration area, integration lock
• peak separation, peak starts and ends
• Store to template method
• parameters will be applied to newly acquired
  chromatograms
• Manual chromatogram modifications
• only actual chromatogram is affected
• store to template method
• copy to other chromatograms
• batch reprocess

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Basic integration parameters
Integration interval
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Integration parameters
Chromatogram method
Integration table individual for each detector
Template method
All operations editable and programmble in time
(including Peak Width and Threshold)
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Toolbars
Autointegration tools Global Peak Width Global
Threshold
Integration Tools integration parameters
settings
Peak Tools peak operations
The crossing of chromatogram and baseline is not
allowed. In such case, the peak start or end is
moved automatically
Baseline Tools baseline modification
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Integration Algorithm
The crossing of chromatogram and baseline is not
allowed. In such case, the peak start or end is
moved automatically
Peak Operations A Peak Apex B
operation Interval Operations Whole peak must be
inside the A-B interval
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Integration Tools
Detector delay 1) first click peak on active
signal 2) then click corresponding peak on first
signal
Rejections area, height, peak width
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Peak Tools
Groups several peaks can be quantitated as
single compound
Solvent Peak not included in result table
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Baseline Tools
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Rejection and Separation

• Rejection
• limits for rejection of already integrated peaks
• peaks bellow set limits could not be added
  manually
• Separation
• functions only from menu or integration table
• Valley to Valley Slope
• sensitivity for automatic Valley to Valley
  separation
• Tangent Area Ratio, Tangent Slope Ratio
• sensitivity for automatic tangent separation
• default values baseline separation

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Storing to Template method

• Save as Template
• use parameters from current chromatogram to new
  chromatogram
• actual method could not be overwritten

• Set Model/Copy from Model
• copy parameters from actual chromatogram to other
  chromatograms

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Integration results

• Integration results are peak retention times,
  areas and heights
• Calibration must be constructed to assign
  compound names and concentrations

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Calibration

• Relationship between retention time and compound
  identity
• Global calibration table
• Identification windows
• RT area, in which peak is identified as a given
  compound
• Reference peaks
• the biggest peak is identified in identification
  window (instead of nearest), identification
  windows for other peaks are shifted accordingly

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Calibration

• Relationship between response and compound
  quantity
• Response
• height or area
• Calibration curve fit
• point to point-linear-quadratic-cubic-sigmoid
• Zero usage
• not used-used as a point-forced through zero
• Recalibration
• replace-weighed average-average
• Response weighing
• none-1/amount-1/response-1/amount2-1/response2

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Calibration window
Transfer data from chromatogram to calibration
Open Calibration
Set level
Active signal selection
Calibration options
Open Standard
Use compound on active signal
Global calibration table
Individual compounds tabs
Current level table
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Calibration options
Calibrate new level for each opened
chromatogram Recalibrate current level
calculation mode
Units used in the result table
Recalibration mode
Enable editing of responses
Actualization of retention times during
recalibrations
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Calibration options
Default response base
Calibration Curve fit type
Response weighting
Zero usage
Set for all compounds in calibration table for
current signal
Indentification windows
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Compound table
Use point
Curve fit Free calibration Point to point
Linear Quadratic Cubic Sigmoid
Response base setting
Compound type Odinary Reference Internal
Standard Reference ISTD
Zero usage Ignore Origin, Compute with
Origin, Curve passes through Origin
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Creating calibration file

• Create new calibration in Calibration window
• command menu File - New
• Set calibration options
• command menu Calibration - Options or icon
• Open integrated chromatogram of a standard
• command menu File - Open primary or icon
• Transfer data from chromatogram to a calibration
  file
• command menu Calibration - All or icon

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Creating calibration file

• Identify peaks
• fill in the peak names and concentrations for
  first level
• Repeat for all standard levels
• Check calibration curves for all compounds
• click on compound tab
• set options (curve fit, zero usage, exclude
  points)
• Save calibration
• command menu File - Save

34
Calibrated calculations

• Linked calibration
• Calibration file is common to all files linked to
  it and all changes in calibration are immediately
  progressed to all opened linked files
• Copy of actual calibration state is stored within
  chromatogram, when saved
• Calibration selection
• Calibration file and calculation type is selected
  in method (template or chromatogram)
• Those parameters should be set in template method
  before starting data acquisition

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Repeated use of calibration

• Each series of samples should be linked to an
  unique calibration file
• Create copy of calibration using Save as or
  Clone
• Clone creates a copy linked to the method
• Clear responses option is available
• Acquire standards
• Recalibrate (by automatic, recalibrate, replace)
• Acquire samples (with linked actual calibration)

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Template method settings
Jump to the linked calibration
Calibration file selection
Calculation type selection
Result table display options
Create and attach copy of calibration file to
template method (option Clear responses)
Calculation Tab
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Chromatogram settings
Select a calibration file
Select the calculation type
Jump to the linked calibration
Response base indication
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Calculation types

• UNCAL
• uncalibrated, only areas and heights
• ESTD
• external standard, most common
• ISTD
• internal standard, calculated from the ratio of
  ISTD and unknown compound
• A) same amount of ISTD in samples and standards
• Internal standard must not be calibrated
• ISTD amount must be set to zero
• B) different amount of ISTD in samples and
  standards
• Internal standard must be calibrated
• ISTD amount must be given
• When those conditions are not met, no results are
  calculated

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Results display
Peak type Error calibration curve could not be
used Free when response factor and Free
Calibration is used
Calculation type If the conditions are not met,
no results are displayed, the reason indicated in
table header
Amount in a) base Total Amount b) base Sample
Amount
Amount calculated from calibration curve
Response and response base
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Printing of results
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Report