Overview. Experiment Specifications. This tutorial will enable you to

Transcription

1 Defining a protocol in BioAssay Overview BioAssay provides an interface to store, manipulate, and retrieve biological assay data. The application allows users to define customized protocol tables representing the workflow of experiments. Tables can contain raw data values, automated calculations, plate inventory information, or a combination of these. A protocol s structure is reflected in a set of tables in BioAssay called a protocol. Once all protocol tables and templates are defined, users can easily import, manipulate, and store new data. There are four major steps in defining a protocol structure and viewing data in BioAssay: 1. Define custom experimental settings. 2. Create table definitions representing the experiment work flow. 3. Define how data will be imported. 4. Import, view and plot data in tables. This tutorial presents the specifications for an example experiment and steps you through the process of defining a protocol in BioAssay. The resulting protocol definition is used to import, store, and view sample data. This tutorial will enable you to Interpret experimental specifications as BioAssay protocol tables Create a plate format from the description of a plate. Construct a table structure for a protocol Use and understand the calculations wizard Define, use and understand file import and graph templates Import and view protocol data from within BioAssay Experiment Specifications The following specifications have been designed for the purposes of this example. The example was designed to be as realistic, yet as general as possible. A lab is running a screening for which they would like to calculate the % inhibition value for a range of drug candidates. Plates contain 6 positive control wells and 6 negative control wells. 1 Defining a protocol in BioAssay

2 Plate Format Plates are described as having 96 wells in all (8 rows and 12 columns. Of the 96 Wells, positive control wells are A1, B1, C1, F2, G2, and H2. Negative control wells are A2, B2, C2, F1, G1, and H1. Wells D1, E1, D2, and E2 are left empty. The remaining wells will contain a sample. Calculations Tests are run and the results are recorded by the instrument taking the measurements in a data file. The resulting calculation desired is % inhibition: % inhibition = 100*(Value-neg)/(pos-neg) Value = raw value neg - negative control average pos - positive control average Data File Format The data file created by the measuring instrument contains raw data for each test instance. The file starts with multiple header lines describing test conditions. Following the header lines is data arranged in a plate format. That is, the data is arranged in a 8 row, 12 column grid with each value placed in the data file grid where the well is found in the plate. A sample data file can be found on the installation CD, or on the client at <BioAssayroot>/Tutorial Assay. The file is called sample_data_for_tutorial.txt. Defining Plate Settings Plate settings describe plate characteristics such as plate size, well distribution, etc. Many plate settings could be defined for this protocol, but there are three which must be defined before proceeding: Physical Plate Type Well Formats Plate Format Physical Plate Type Physical plate types define the layout of a plate. That is, the number of wells and how those wells are arranged in the plate. There is a default Physical Plate Type that fits the plate described for this protocol, called 96 Well Plate. The 96 Well Plate can be pictured as an 8x12 grid where each cell in the grid represents a well in the plate. Therefore, an additional physical plate format will not have to be defined. Well Formats Well Formats define the general contents in a well. In other words, the well format describes the contents of the well without indicating the actual substance inside the well. The following default well formats are sufficient for the purposes of this protocol: Compound - can contain a sample Empty - will remain empty Positive Control - will contain a positive control Negative Control - will contain a negative control Defining a protocol in BioAssay 2

3 Plate Format Plate Formats define which wells in a plate (with a given physical plate type) contain particular well formats. There are no default Plate Formats defined that will fulfill the requirements of this protocol. The Plate Format that we define will take into account all of the plate format specifications described under Plate Format on page 3. The specifications tell us what type of substance is contained in each of the 96 wells in the plate. To define a new plate format for this protocol: 1. After logging in, expand Settings > Plate Setup. 2. Click on the Plate Formats folder. A table defining all available plate formats appears in the right frame. 3. Click Edit. 4. Click Add. A new row appears. 5. Enter Tutorial Format as the name of the new format and select the 96 Well Plate from the Physical Plate Type listbox. 6. Click Save. 7. Double click on the row defining the new plate format (Tutorial Format). A grid of 96 cells (each representing a well) appears in the right frame. The default is a well format of Compound in each cell. Next, we will have to assign a well format to each cell. 8. Click Edit. 9. Populate the appropriate cells with the Compound well format. a. Highlight cell D1. b. Select Empty from the pull-down menu. c. Click off of cell D1. d. Highlight cell D1 again e. Select Copy from the Actions menu. f. Highlight each of the following cells (one by one) and select Paste from the Actions menu: D2, E1, and E Populate the appropriate cells with the Positive Control well format. a. Highlight cell A1. b. Select Positive Control from the pull-down menu. c. Click off of cell A1. d. Highlight cell A1 again. e. Select Copy from the Actions menu. 3 Defining a protocol in BioAssay

4 f. Highlight each of the following cells (one by one) and select Paste from the Actions menu: B1, C1, F2, G2, and H Populate the appropriate cells with the Negative Control well format. a. Highlight cell A2. b. Select Negative Control from the pull-down menu. c. Click off of cell A2. d. Highlight cell A2 again. e. Select Copy from the Actions menu. f. Highlight each of the following cells (one by one) and select Paste from the Actions menu: B2, C2, F1, G1, and H1. The grid should now look like this: Defining the Protocol There are many ways to define a protocol. This is a plate based protocol, so for the purposes of this tutorial, we have chosen to define this protocol with two tables: a Plates table and a Wells table. This is not the only way to define this protocol and you should feel free defining protocols in a way that makes the most sense to you. Raw data recorded from each well will be contained in the Wells table. Several rows in the Wells table are linked to one row in the plates table to indicate which wells exist in a particular plate. Therefore, calculations summarizing well data from an entire plate are found in the Plates table. Creating the Protocol Before defining tables for the protocol, first create a new protocol in the Protocols folder in the navigator. To create the new protocol: 1. Expand the Protocols Folder in the navigator. 2. Browse to the project to which the new protocol belongs. NOTE: See the BioAssay user s guide for more details about folders and projects in the navigator. 12. Click Save to save the changes. The resulting grid (pictured above) represents the plates described in the protocol specifications. When new plates are created, this plate format will be used. For more information about creating new plates, please see Creating a Plate (the next section). 3. Right-click on the project icon and select New Protocol. 4. Enter a name (Tutorial Assay) and click OK. Defining a protocol in BioAssay 4

5 A window allowing you to select a template appears. The protocol definition appears. Templates set up a shell for a protocol definition. There are templates for plate based protocols as well as non-plate based protocols. 5. For this protocol, highlight the Initial Screen Protocol Template. NOTE: For the purposes of this tutorial, we will NOT be creating an inventory linked protocol. For more information about inventory linked protocols, please see the BioAssay User s Guide or contact CambridgeSoft s technical support. 6. Click OK. A new protocol is inserted under the appropriate project in the navigator. Protocol Definition Because we used the Initial Screen template, the tables for the protocol have already been preliminarily defined. To view the new protocol s definition: 1. Browse to the Tutorial Assay protocol in the navigator. 2. Double-click the Tutorial Assay protocol. 3. Select the Definition Tab. Wells Table The Wells table will contain data concerning one well. The template used inserts the following fields in the Wells table: Well ID - an auto generated ID identifying the well. Well Format Compound ID - the compound ID, helps to identify what compound is in which well Concentration (um) - concentration of the substance in the well Raw Value - raw data for each well Plates Table The Plates table summarizes the wells table data. This table holds information pertaining to an entire plate. The template used inserts the following fields in the Plates table: Plate ID - an ID for the plate Plate Format - the plate format used for the plate (see Plate Format on page 3 for more information) Pos control average 5 Defining a protocol in BioAssay

6 Neg control average NOTE: Calculation fields are identified by the icon in the Protocol Tree. These fields are assigned the calculation field type, but the calculation is not defined until you explicitly define it. See Defining the Calculations with the Calculation Wizard on page 6 for more information about defining the calculations. Import Sessions Table The import sessions table hold data summarizing a number of plates, all imported at the same time. The template used inserts the following fields in the Import Sessions table: Import Date Scientist Import Comment Adding additional fields For this simple protocol, we only need to add one field to our protocol definition in order to complete the tables definitions: the % inhibition. The % inhibition is a value calculated for each well, so this field will be found in the Wells Table. For information about adding the % inhibition field, please see Percent Inhibition on page Click Edit Calculation The Calculation Wizard appears. 3. Highlight Average under the Aggregate Tab. 4. Click Next. 5. Select Wells Table from the Table listbox and Raw Value from the Field listbox. 6. Click Next. 7. Click Add (to add a condition). 8. Select Wells Table from the Table listbox and Raw Value from the Field listbox. 9. Click Next. 10. Click Add (to add a condition). 11. Select Well Format from the Field listbox, is from the Is listbox. 12. Double-click the Value textbox. 13. Select Constant Value and enter Positive Control. Defining the Calculations with the Calculation Wizard Positive Control Average Take the following steps to define the positive control average with the Calculation Wizard: 1. Click on the Pos Ctl Average field under the Plates table in the Protocol Definition Tree. The field definition appears in the right frame. 14. Click OK. 15. Click Next. 16. Click Finish. Defining a protocol in BioAssay 6

7 Negative Control Average Take the following steps to define the negative control average with the Calculation Wizard: 1. Click on the Neg Ctl Average field under the Plates table in the Protocol Definition Tree. The field definition appears in the right frame. 2. Click Edit Calculation The Calculation Wizard appears. 3. Highlight Average under the Aggregate Tab. 4. Click Next. 5. Select Wells Table from the Table listbox and Raw Value from the Field listbox. 6. Click Next. 7. Click Add (to add a condition). 8. Select Well Format from the Field listbox, is from the Is listbox. 9. Double-click the Value textbox. 10. Select Constant Value and enter Negative Control. 11. Click OK. 12. Click Next. 13. Click Finish. Percent Inhibition To add the % inhibition field to the Wells Table: 1. Click on the Wells Table in the Protocol Definition Tree. 2. Right-click Wells in the Protocol Definition Tree and select New Field. The field tab opens with default values and the new field is added to the Protocol Definition Tree. 3. Type % inhibition in the Field Name listbox. 4. Select Calculation from the Field Type listbox. 5. Click Save Changes. Take the following steps to define the % inhibition with the Calculation Wizard: 1. Click on the % inhibition field under the Wells table in the Protocol Definition Tree. The field definition appears in the right frame. 2. Click Edit Calculation. The Calculation Wizard appears. 3. Click on the Mathematical Tab and highlight % Inhibition. 4. Click Next. 5. Create the following table: 6. Click Next. 7. Click Finish. 7 Defining a protocol in BioAssay

8 Importing Data Data is imported into BioAssay through the use of File Import Templates. Data import templates inform the application where to find particular data and how to populate the defined protocol tables. Creating a File Import Template The BioAssay application provides a wizard to help users define file import templates. To define a file import template for the sample data file: 1. Click on the File Import Templates folder in the Protocol Definitions Tree. 2. Right-click and select New Template. The File Import Wizard appears. 3. Enter Sample Data File in the Template Name textbox. 4. Enter a description in the Description textbox. 5. Click Next. 6. The file is arranged in a Plate Block, so click Plate Block. 7. Select Tabs from the delimiter listbox. If you look at the file in Excel, you will see that the following is the proper number of lines for the Plate Diagram File setup: NOTE: Sample data is found on the BioAssay CD and <BioAssayroot>/Tutorial Assay on the client. The file is called sample_data_for_tutorial.txt. 8. Click Next. The Map Fields screen appears. 9. Fill in where the data should be imported from. a. Select Plate Block Data from the listbox next to Raw Value (this indicates the plate block data should be imported into the Raw Value field). b. Select Prompt Me from the listbox next to Import Date, Scientist, and Import Comment (this indicates the user should be prompted during import for values for these fields). c. Select Block Header Cell from the listbox next to Plate ID. d. Enter 2,1 in the far right column next to Plate ID. This indicates that the Plate ID is found in the second row, first column of the plate header. Defining a protocol in BioAssay 8

9 e. Select Use Default from the list box next to Plate Format. f. Select the Tutorial Format in the far right column next to Plate Format (this indicates the Tutorial Format will always be used when plates are imported using this file import template). 10. Click Finish. Using the Import Data Wizard After a file import template is defined, data is imported through the use of the data import wizard. The Data Import Wizard requests a filename and a file import template to use and then imports data according to the parameters set in the file import template. To import the data in our sample data file: 1. Click on the Tasks Tab for the Tutorial Assay. 2. Select the Import a Data File link. 3. Select Sample Data File from the listbox. 4. Click Next. 5. Click on Browse and select the sample data file. NOTE: Sample data is found on the BioAssay CD and <BioAssayroot>/Tutorial Assay on the client. The file is called sample_data_for_tutorial.txt. 6. Click Next. 7. Enter data for any fields that you are prompted for. If you followed the above instructions, you should be asked for the following data: Import Date Scientist Import Comment 8. Click OK. 9. Click Next. 10. Click Finish. If the option to show the data before loading was left selected in the wizard, a table displaying the data to be loaded appears. CLick Load File to continue. 11. Click on the Data Tab. A search screen appears. 12. Click Retrieve All. The Import Sessions table appears. Drill down into the Plates and Wells tables by clicking on the plus sign in the left most column of the data table. 9 Defining a protocol in BioAssay

10 Defining a protocol in BioAssay 10