Validation Sample Set Generator
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1 Validation Sample Set Generator General Information This tool is an example program which demonstrates some capabilities of the Empower Toolkit. It was not created using Waters Software Development Lifecycle. This tool was tested in a limited fashion and is not structurally validated by Waters Corporation. This tool is open source and Waters freely distributes the source code for use and modification by customers that have purchased the Empower Toolkit Option. This source code was created using Visual Basic version 6.0. For more information on use of the Toolkit, see the documentation provided with the Empower 2 Toolkit option. The Validation Sample Set Generator (VSG) tool is intended to be used in conjunction with the Empower 2 Method Validation Manager option to create the necessary instrument methods, method sets, and sample set methods to satisfy the robustness experimental plan specified for a particular robustness test in an MVM validation protocol method. The definition of robustness of an analytical procedure is the measure of its capacity to remain unaffected by small, but deliberate, variations in method parameters and provides an indication of its reliability during normal usage. Hence, in a robustness experiment the user deliberately varies specified parameters, referred to as factors. The variation is performed at a high value, a low value and a centerpoint, or nominal value (the normal value specified by the method you are validating). These factors can include sample and reagent preparation variations such as sonication time, shaking time, extraction time, sample ph, and so on. These factors can also include instrument method variations such as column temperature, sample compartment temperature, flow rate, mobile phase composition, detector wavelength, and so on. The Empower 2 Method Validation Manager (MVM) software provides a robustness experimental plan, based on user input of what factors are to be varied and what type of plan to be used. This experimental plan dictates the combination of factor variations (experiments) necessary for a set of injections (i.e. a sample set) on an injection-byinjection basis; an example experimental plan is shown below. The factor values are represented either by -1 (the low value of the factor) and +1 (the high value of the factor), or by specifying the factor and values explicitly (both representations are available in MVM). The VSG tool requires user inputs to specify which factors map to sample, preparation, or instrument variations. The VSG tool also requires a sample set method which specifies the method set containing the nominal instrument method. The tool then translates the experimental plan into the necessary instrument methods, method sets and sample set(s). Page 1 of 10
2 ½ Fraction (2 4-1 ) Experimental Plan example (without centerpoints): Experiment Factor 1 Factor 2 Factor 3 Factor This tool was not developed using Waters software development lifecycle and has not been structurally validated. It is the user s responsibility to validate this tool, if validation is necessary. Hardware and Software Requirements The Validation Sample Set Generator tool supports the same computer hardware/software configuration requirements as the base Empower 2 application. The Validation Sample Set Generator tool supports the following chromatographic systems: Alliance (at firmware supported by Empower 2) ACQUITY Binary Solvent Manager, Sample Manager, Column Heater, Tunable UV + Photodiode Array (at firmware supported by Empower 2) A1100 Isocratic Pump, Quaternary Pump, Binary Pump, Auto Injector, Auto Injector With Heater, VW Detector, RI Detector, Column heater, MW Detector, DA Detector (at firmware supported by Empower 2) Installation of the Validation Sample Set Generator Installing the Validation Sample Set Generator requires the use of a privileged account with permission to write to the system resources. Please ensure that the account used to perform the installation has sufficient privileges. Note that a Domain Administrator account may not have the same privileges or permissions as the Local Administrator account. Page 2 of 10
3 1. Navigate to and double-click on the "Setup.exe" file. 2. Follow all prompts. If you receive the following message during installation of this tool, select Yes. Use of the Validation Sample Set Generator Use the Start menu to navigate to the Validation Sample Set Generator. (For example, Start > All Programs > Empower > Validation Sample Set Generator.) Login using the following login dialog: Page 3 of 10
4 The following describes the different fields used by the Validation Sample Set Generator to determine how to create the required sample sets. Refer to Figure 1: Validation Sample Set Generator User Interface. Creation Parameters Validation Method Robustness Test No. of Preparations Use a unique vial for each line Use this dropdown list to specify the MVM validation protocol method which contains the desired robustness test. Use this dropdown list to specify the MVM robustness test for which you want the Validation Sample Set Generator to automatically create instrument methods and sample set methods. Use this selection to specify how many sample preparations will be performed for each sample (experiment). When you specify a number of Preparations greater than 1, it is recommended to also select Use a unique vial for each line so that each sample (experiment) in the resulting sample set method(s) will be duplicated the specified number of times. Note: This setting does not affect the Sample Preparation field in the resulting sample set method. The user will need to manually specify the Sample Preparation value for each line in the sample set method. By selecting this you turn off intelligent vial allocation. Every new sample set line will use its own unique vial (and so be assigned a unique vial location depending upon your current Page 4 of 10
5 No. of Injections Include Standards No. of Centerpoints Template Sample Set Method instrument plate). This selection specifies the number of injections for each sample (experiment) in the resulting sample set method(s). This selection specifies that the Validation Sample Set Generator should place standard injections in the resulting sample set method(s). These standards will be created using perturbations of the factors defined as Instrument Factor Types but will hold all other Factor Types at the Nominal value. (If you need to run standards using perturbations of all factor types, you will need to manually enter standards in the resulting sample set method. This can be easily accomplished by highlighting an appropriate sample row and selecting Edit > Insert Row.) Note: The Robustness test checkbox field will be enabled for any standard lines in the resulting sample set method. This check box will need to be manually disabled. This selection specifies that the Validation Sample Set Generator should place the specified number of Centerpoint injections in the resulting sample set method(s). The centerpoint injections are run at the nominal factor conditions and are placed in appropriate locations in the resulting sample set method(s). The position is determined using the same criteria as standard lines, this is based upon the instrument method factors equilibration time. Note: The Sample Preparation field for the centerpoint injections in the resulting sample set method(s) will be specified as Reference. This field will need to be set appropriately (to Preparation 1, Preparation 2, and so on.) The first line of the template SampleSet Method is used as the template for all lines to be created. No other lines are examined/required within the template sample set method. All of the parameters in the template sample set line are copied exactly unless they are altered by a validation factor or they represent a key field e.g Sample Name. The first line in the template sample set method must contain a valid method set which specifies an appropriate instrument method. This instrument method should contain the nominal instrument parameters (the nominal parameter settings per the method that you are validating) and will be used as the starting point by the Validation Sample Set Generator tool. When instrument factors require changes, in order to affect these changes new instrument methods must be created. For each instrument factor change the tool will create a new Page 5 of 10
6 Reset instrument method and a new Method Set containing this instrument method. The Method set will then be assigned to the appropriate Sample Set Line. Instrument methods and method sets are reused where possible. Selecting this button clears the following fields so that you can initiate the Validation Sample Set Generator process with a different robustness test: Validation Method Robustness Test Template Sample Set Method Vial Information Maximum Extraction Volume from each Vial (ul) Sample Sets have unique vials Sample Sets share vials Specify the maximum volume that can be injected from each vial so that the Validation Sample set Generator knows when to specify a new vial for the same sample. (Other factors such as vial dead volume, needle overfill volume, and syringe volume also need to be considered and subtracted from the total available volume.) This selection specifies that if multiple sample set methods are created, the sample vials in one sample set method are not reused in the other sample set method(s). This selection specifies that if multiple sample set methods are created, the sample vials in one sample set method are to be reused in the sample set method(s). Method Names to Use Sample Sets Method Sets Instrument Methods Add method type prefixes (SS, MS, IM) Specify the root name of the resulting sample set method(s) using alphanumeric characters. Do not use special characters such as /, %, +, #, and so on. Specify the root name of the resulting method set(s) using alphanumeric characters. Do not use special characters such as /, %, +, #, and so on. Specify the root name of the resulting instrument method(s) using alphanumeric characters. Do not use special characters such as /, %, +, #, and so on. Specify that the resulting methods should contain an identifying prefix: SS = sample set method MS = method set IM = instrument method Note: The resulting method will also contain an incrementing numeric suffix. Keep in mind that the maximum number of Page 6 of 10
7 characters that can be used in a method name is limited to 30 characters in Empower 2. Factor Definitions Factor Name Factor Type The Factor Name (custom field) specified in the MVM robustness acquisition parameters. Use this selection when the factor is a sample set Sample Set parameter (for example, Analyst). The factor values in the corresponding field in the sample set method will be set according to the robustness experimental plan in Method Validation Manager. Instrument Use this selection when the factor is an instrument method parameter, for example Flow Rate. The Validation Sample Set generator will modify the appropriate instrument methods according to the robustness experimental plan in Method Validation Manager and specify them in the appropriate sample lines in the sample set method(s). The factor values in the corresponding field in the sample set method will be set according to the robustness experimental plan in Method Validation Manager. Ignore Preparation Custom The following fields will be enabled and must be set appropriately: Instrument Type Instrument Method Parameter Relative Equilibration Time Use this selection when a factor in the MVM robustness parameters should be ignored. Use this selection when the factor is a sample preparation parameter, for example Sonication Time of sample. The Validation Sample Set generator will change to a new sample vial in the resulting sample set for each factor perturbation. The User Set Up Information documents a mapping of which vial numbers are set to which factor perturbations in the resulting sample set method(s). The factor values in the corresponding field in the sample set method will be set according to the robustness experimental plan in Method Validation Manager. Use this selection when you will manually set up this factor in the resulting instrument methods and sample set methods. After the sample sets have been created an information box displays which sample set lines (and if relevant, Page 7 of 10
8 instrument methods) are associated with each factor setting. The display allows the user to manually assign the correct values to sample set lines and instrument methods. For example, this tool does not have the capability to set changing gradient percentages or slopes. These settings can be manually changed by the user once the Validation Sample Set Generator creates the sample set method(s). Minimum Value Nominal Value Maximum Value Requires Own Sample Set Generate Instrument Method Instrument Type Instrument Method Parameter Relative Equilibration Time The relevant factors values will be specified in each sample set line. The Lower Value (-1) specified in the MVM robustness acquisition parameters. The Nominal Value (0) specified in the MVM robustness acquisition parameters. The Upper Value (+1) specified in the MVM robustness acquisition parameters. For use with Sample Set, Instrument, Preparation and Custom Factor Types. Specifies that when the factor is varied, a new sample set method must be created. (For example, a factor of Analyst would require each perturbation of Analyst to be placed in a separate sample set method.) For use with a Custom Factor Type only. Specifies that a separate instrument method should be created for each perturbation of this factor. It is up to the user, however, to set the corresponding parameter in the resulting instrument method. For use with an Instrument Factor Type only. Select the instrument to which the factor applies. For use with an Instrument Factor Type only. Select the instrument parameter to which the factor applies. For use with an Instrument Factor Type only. Select from: N/A Short Medium Long For any factors which will require equilibration in between injections, this setting allows you to specify the equilibration time relative to the other factors which also require an equilibration time. This setting affects the order of instrument methods in the resulting sample set method(s). For example, if you are using 2 factors which require an equilibration time such as column temperature and flow rate, the temperature change requires a longer equilibration time than the flow rate change. Hence, the oven factor should be set as Long and the flow rate should be set as Short. The Validation Sample Set Generator tool will attempt to order the sample set method lines so that they are ordered from the lowest to the highest Page 8 of 10
9 oven temperature. Secondarily, the tool will attempt to order the sample set method lines from the lowest to the highest flow. Once the sample set method(s) is created, the user must add the appropriate Equilibration lines. (Select a row and choose Edit > Insert Row. Set the Function field to Equilibrate and the Run Time equal to the time required for the equilibration. Note: This functionality works best when the factor values are numeric, rather than alpha-numeric strings. Figure 1: Validation Sample Set Generator User Interface After specifying the required information, click the Create SampleSets button. When the sample set methods are complete, a SampleSets Created Successfully message is displayed. Upon successful completion, the User Setup Information window will be displayed. This window provides a mapping of the Preparation type factors to the vials listed in the sample set method(s) as well as a mapping of the custom field value to each sample set line. In the example User Setup Information below, Sonication Time has been set as a Preparation factor type so a mapping of the appropriate factor value to use for each vial is Page 9 of 10
10 provided. Additionally, StrokeVol was set as a custom factor type, so a mapping of the custom field value to each sample set line is displayed with the name of the relevant line and the name of the method set that contains the instrument parameters for this line. Example User Setup Information: **SS_Alpha_1** SS_Alpha_1 : VIAL INFORMATION Vial Id : 1 Sonication time: 15 Vial Id : 2 Sonication time: 10 Vial Id : 3 Sonication time: 20 SS_Alpha_1 : CUSTOM FIELD INFORMATION Line Name = Sample0 - StrokeVol : 50, Method Set Name : MS_Alpha_2 Line Name = Sample1 - StrokeVol : 130, Method Set Name : MS_Alpha_2 Line Name = Sample2 - StrokeVol : 50, Method Set Name : MS_Alpha_2 Line Name = Sample3 - StrokeVol : 130, Method Set Name : MS_Alpha_2 Line Name = Sample4 - StrokeVol : 50, Method Set Name : MS_Alpha_2 Line Name = Sample5 - StrokeVol : 130, Method Set Name : MS_Alpha_2 Line Name = Sample6 - StrokeVol : 50, Method Set Name : MS_Alpha_2 Line Name = Sample7 - StrokeVol : 130, Method Set Name : MS_Alpha_2 Line Name = Centerpoint - StrokeVol : 100, Method Set Name : MS_Alpha_3 Line Name = Sample8 - StrokeVol : 50, Method Set Name : MS_Alpha_1 Line Name = Sample9 - StrokeVol : 130, Method Set Name : MS_Alpha_1 Line Name = Sample10 - StrokeVol : 50, Method Set Name : MS_Alpha_1 Line Name = Sample11 - StrokeVol : 130, Method Set Name : MS_Alpha_1 Line Name = Sample12 - StrokeVol : 50, Method Set Name : MS_Alpha_1 Line Name = Sample13 - StrokeVol : 130, Method Set Name : MS_Alpha_1 Line Name = Sample14 - StrokeVol : 50, Method Set Name : MS_Alpha_1 Line Name = Sample15 - StrokeVol : 130, Method Set Name : MS_Alpha_1 If further modifications of the sample set method(s) are required, you can do so in the Sample Set Editor. To add lines, select an appropriate row and select Edit > insert row. To re-order lines, use drag and drop functionality. Page 10 of 10
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