Академический Документы
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Культура Документы
January 2014
Janick Mathys
VIB-BITS
Contents
1.
Installation ....................................................................................................................................... 5
2.
Documentation................................................................................................................................ 5
3.
3.2.
3.2.1.
Families ............................................................................................................................ 7
3.3.
3.4.
3.5.
Import data...................................................................................................................................... 9
5.1. Data table types ......................................................................................................................... 10
5.1.1. XY tables .............................................................................................................................. 10
5.1.2. Column tables ...................................................................................................................... 10
5.1.3. Grouped tables .................................................................................................................... 11
5.1.4. Parts of whole tables ........................................................................................................... 11
5.1.5. Contingency tables .............................................................................................................. 11
5.1.6. Survival tables...................................................................................................................... 12
5.2. Limits of data tables ................................................................................................................... 12
5.3. Data types................................................................................................................................... 13
5.4. Missing values ............................................................................................................................ 13
5.5. Import data................................................................................................................................. 13
5.5.1. Manually enter data in Prism .............................................................................................. 14
5.5.2. Copy from another Prism table and paste .......................................................................... 14
5.5.3. Copy from an Excel table and paste .................................................................................... 15
5.5.4. Import data files .................................................................................................................. 15
5.5.6. Create X-values of XY tables automatically ......................................................................... 18
5.6. Open existing Prism Projects ...................................................................................................... 18
5.7. Change data tables ..................................................................................................................... 19
5.7.1. Change table format ............................................................................................................ 19
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1. Installation
BITS offers PRISM licenses for free to VIB scientists. You can find the installation procedure on our
website, you have to log on using your VIB account (the account you use to check your VIB mail):
http://www.bits.vib.be/index.php/software-overview/graphpad-prism
2. Documentation
The software runs on Windows and Mac. You can find a lot of documentation online:
- Prism user guide: http://www.graphpad.com/guides/prism/6/user-guide
- Statistics guide: http://www.graphpad.com/guides/prism/6/statistics/
- Curve fitting guide: http://www.graphpad.com/guides/prism/6/curve-fitting/
You can easily access these tutorials within Prism by expanding the Help section in the top menu.
3. User interface
Video tutorial on Prism user interface: http://www.bits.vib.be/index.php/training/131-basic-statistics
3.1.
When you start Prism, the Welcome dialog is opened, containing the following sections:
Red: select one of the six types of data tables that Prism supports (see section 5.1 for a description of
these table types). When you make a selection a new table of this type is generated.
Orange: open existing projects (see section 5.6), example graphs or clone graphs (see section 8.4).
Green: more info on the selected table type: description and an example of a typical graph.
Purple: set the details of the new table: do you want an empty table to import your own data in
(upper part) or do you want to work with the example data in Prism (lower part)?
In the former case you have to specify the format of the data you want to enter. In the latter case
you have to select the type of analysis that you want to perform. Each table type has a list of analyses
that can be performed on it (see appendix for an overview). Selecting an analysis type fills the table
with example data that are ideal for doing that kind of analysis but the analysis itself is not
performed: you have to do the analysis yourself.
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For discussing the user interface we will open the example column table containing data for an
unpaired t-test. Prism will load the Ozone correlations project.
3.2.
Allows users to browse to the information they want to work on. The navigator shows your data, and
analysis results using a specific hierarchy:
Projects contain:
Each of these five folders can contain multiple pages, called sheets. The navigator shows all the
sheets of the project that is currently open.
3.2.1. Families
Prism remembers the logical links between sheets. When you edit or replace data, Prism
automatically re-computes linked analyses and redraws linked graphs. A group of linked sheets is
called a family. Families are shown at the top of the navigator: e.g. when you open the sample XY
data table for correlation you see Family of Ozone correlations at the top of the navigator. The
names of the sheets that are part of the family are shown in bold in the navigator. You can also
expand the family to see which sheets belong to it. As said before Prism creates families
automatically by keeping track of the links between sheets but you can also create, delete or move
families yourself: http://www.graphpad.com/guides/prism/6/user-guide/families_of_sheets.htm
Families allow you to easily repeat your analysis with new data by duplicating the family. To duplicate
a family you can choose one of these three strategies:
Prism duplicates the data table and all linked graphs and result tables. If you replace the data on the
new data table, all analyses and graphs will be updated.
Prism will ask you to assign a name to the new (duplicated) data table. Suppose that the data table is
"Ozone" and is linked to "Ozone correlations". If you duplicate the family, you can enter "CO2" as the
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name of the new data table. Prism will then rename the linked graph to "CO2 correlations"
automatically. It can only do this because the exact name of the data table is part of the graph name.
Note the distinction between duplicating a sheet and duplicating a family. If you duplicate a graph,
the copy will link to the same data table as the original. This is useful if you want to make two graphs
of the same data. But if you want to make two similar graphs of different data, you have to duplicate
the family or clone the graph (see section 8.4).
3.3.
Shows the sheets that you open by simply clicking the name of the sheet in the navigator.
3.4.
Bottom toolbar
You can also open an overview of all sheets of one of the five components by clicking the name of a
sheet/folder in the navigator and click Gallery (
) in the bottom toolbar. This will open a gallery
view in the Main window where you see all sheets of that component, e.g. all graphs of your data.
To remove the gallery and return to the normal sheet view, click
You can hide the navigator by clicking Hide / Display (
again.
3.5.
The top menu and the upper toolbar give access to all the actions that you can perform in Prism.
Prism is very redundant: each action can be done in multiple ways and the top menu and the upper
toolbar contain more or less the same features. For instance if you make a mistake, you can always
undo your last moves by following one of these strategies:
Edit in Top menu -> Undo
Undo (
Both the top menu and the upper toolbar are divided into sections and although the arrangement of
these sections is different, they do provide the same actions.
The buttons of the upper toolbar and the items in the dropdown menus of the top menu will change
depending on which type of information is open in the main window. When you are looking at a data
table you will see different buttons in the upper toolbar compared to when youre looking at a graph.
Prism (
New (
) in the File section of the upper toolbar -> New Project File
When you create a new Project you are redirected to the Welcome dialog (see section 3.1.) where
you have to select (as always) a data table type. The type that you choose is not important; Once the
project is created you can open all kinds of data in it.
Save (
) in upper toolbar
5. Import data
Once you have created a project, you can load data into the project. Data is entered in a data table
that consists of rows and columns. The columns can be divided into sub-columns that are intended to
hold data from replicate measurements. If you enter replicate values, Prism will automatically
calculate and plot error bars. You can also enter pre-calculated standard deviations or standard
errors in Prism if you have calculated them elsewhere, but there is no need to do so.
There are six types of data tables in Prism (see section 5.1).
The choice of table type is very important: each graph and analysis type is linked to a certain table
type. For graphs Prism is not very stringent: it will allow you to use each graph type on each table
type, but the result doesnt always look good when you combine a graph that is linked to a certain
table type with another type of table. Often Prism struggles with the titles of axes, legends
Prism is more stringent about analyses: analyses are only allowed for specific data table types. If you
don't choose the appropriate type of table, you won't be able to perform the analysis you have in
mind. So choose a table type based on the organization of your data and the analysis you wish to
perform.
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Column X contains X-values, values of the variable that you manipulate (time, temperature,
concentration) and that is used to plot the others against. The other columns (one or more Y
columns) contain the actual measurements (continuous data), e.g. in the example above contains
column A measurements from one group of subjects. Replicates are placed in sub-columns, e.g. in
the example above column A contains 3 sub-columns with replicate measurements for this one
group of subjects. In this way replicate measurements are recognized by Prism and automatically
used for calculating error bars.
There is only one X-column per XY-table but there can be multiple Y columns.
5.1.2. Column tables
In a column table each column contains the measurements for a specific group, so column tables
are also meant to hold continuous data that is grouped according to a single grouping variable.
Prism defines different groups by using separate columns, one for each group. Column A contains
measurements for the group of patients that received placebo; column B for the group that received
active drug.
By nature, column tables contain replicates in each column so column tables do not contain subcolumns. This table can hold paired or unpaired data. If the data is paired then all values coming from
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the same individual are placed in the same row. In example shown above the data cannot be paired
since both columns contain a different number of values.
5.1.3. Grouped tables
Grouped tables contain measurements for two grouping variables.
One grouping variable (Men vs. Women) is defined by the rows; the other grouping variable (Control
vs. Treated) is defined by the columns. Sub-columns contain data from replicate measurements.
5.1.4. Parts of whole tables
A parts of whole table is used to hold categorical data. It is the categorical counterpart of a Colum
table. So instead of measurements the table contains the number of observations that fall into a
certain category. It is essential that you enter the actual number of observations that you counted.
Do not enter normalized values or percentages. Since the values are counts, they cannot be negative
and must be integers. Prism will not let you enter a minus sign or decimal point.
The example below shows the number of students who received grades of A, B, C, etc.
5.1.5. Contingency tables
Contingency tables are the categorical counterpart of Grouped tables. So instead of one variable that
defines the categories, you have two. The categories are defined by the rows and the columns.
In the example above, the two rows represent different treatments (two categories: standard and
experimental treatment), the two columns represent two alternative outcomes (two additional
categories: graft patent and graft obstructed). In total you can make 4 treatment / outcome
combinations, so you have a total of 4 categories.
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Each value in the table is the number of observations for a specific category. As in Parts of whole
tables you have to enter the actual count, not percentages or fractions.
5.1.6. Survival tables
Survival tables contain data for survival analysis, a branch of statistics that studies the occurrence of
events in time. Events are in most cases binary (yes or no) like death, failure, injury, sickness,
recovery from sickness, exceeding a threshold
Survival analysis answers questions like: How many out of 100 people will survive until 86 years?
Whats a persons chance of surviving past 20 years? Are there environmental factor that increase or
decrease the death rate... Therefore, many clinical trial data are analyzed by survival analysis e.g. the
effect of hormone treatment in women on the incidence of coronary heart diseases, the effect of
aspirin treatment on the mortality after myocardial infarctions
Each row represents a distinct subject. Each Y-column represents a treatment. The X-column
contains elapsed time, usually in days. Note that you cannot use dates since Prism does not let you
enter dates. For each subject, you have to enter a code for Y:
Enter 1 for a subject if the event (death) occurred at the time entered into X.
Enter 0 if the data are censored. Data are censored when you simply dont know what
happened to the subject after that time, or you do know but cant use the information.
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This opens a window that is very similar to the Welcome dialog (see section 3.1) where you can
specify the type of data table you want to create.
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First you have to specify if you want to enter new data or work with the built-in example data. For
either choice you have to specify the details of the data. This is exactly the same procedure as
described for the welcome dialog (see section 3.1). Note that for a table in which you are going to
enter data, you have to specify exactly what the data look like: e.g. the number of replicates,
whether you will use pre-computed standard errors or not
There are six ways of entering data into Prism:
1. Manually type in the data values in a table
2. Copy from another Prism table and paste
3. Copy from an Excel table and paste
4. Import an Excel file
5. Import a .txt or .csv file
6. Automatically generate data values according to a mathematical formula
5.5.1. Manually enter data in Prism
Select a cell with the mouse and simply type in the value you want to enter
5.5.2. Copy from another Prism table and paste
You can copy/paste data from one Prism table to another. To copy data from a Prism table, you have
to select the data you want to copy and follow one of the following strategies:
Edit in the Top menu -> Copy
Right click the selection -> Copy
Copy (
Paste the data in the other table by placing your cursor in the cell that will become the upper-left
corner of the pasted data and follow one of the following strategies:
Edit in Top menu -> Paste -> Paste Data / Paste Embed / Paste Link
Right click the cell in the new data table -> Paste -> Paste Data / Paste Embed / Paste Link
Edit in Top menu -> Paste special
Paste (
Paste special (
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The Source tab will let you choose whether to embed or link to the original file (Windows only):
Insert data only will not keep a link to the original file
Insert and maintain link will keep the link
If you choose to keep a link and you change something in the original Excel file, the change will
become apparent in the Prism copy once you have clicked Save in Excel. This is different with paste
linking, where you see the change in the copy table from the moment you have made it in the
original table.
The View tab allows you to preview the result of your data import.
The Filter tab allows to specify the representation of missing values in your data set and to exclude
rows or columns from the import.
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The Placement tab allows to specify the location of the column names and to rearrange the data as
you bring it into Prism.
Transpose means that you switch rows and columns: the data that are located in row 1 will
transfered to column 1 and vice versa and this for all rows and columns:
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If all data are stacked into a single column (or row) and you want to organize the data into multiple
columns you can use the By rows and By columns options.
An example of a data set consisting of a single column:
Aa_replicate1
A
B
Aa_replicate2
->
a
replic1 replic2 replic3 replic1 replic2 replic3
Aa_replicate3
?
b
replic1 replic2 replic3 replic1 replic2 replic3
Ab_replicate1
Ab_replicate2
Ab_replicate3
Ba_replicate1
Ba_replicate2
Ba_replicate3
Bb_replicate1
Bb_replicate2
Bb_replicate3
There are two grouping variables: A and B along the columns and 1, 2, 3, 4 and 5 along the rows.
Each measurement was repeated thrice. To load this data into a Grouped Table with 5 rows, 2
columns and 3 subcolumns each, you can use the following settings:
Import from such files is done in exactly the same manner as from Excel files with the exception that
you have to specify the role of the comma.
Note that you can't open a text file directly from Prism's Welcome dialog or with the File Open
command. To import a file, you must first create a new Prism project (or open an existing one), go to
a data table, position the insertion point, and then choose the Import command.
5.5.6. Create X-values of XY tables automatically
Often the X column of an XY table is a series of values that follow a mathematical formula, e.g.
increasing by 1... To create an X column like this you have to select the cell of the X column where
you want to start the series and choose:
Insert in Top menu -> Create Series
Note that you can use this for any column in any table but in practice you mostly use it for generating
X- values.
After you have used Prism a bit, you may see files listed under the heading Unsaved Files. When you
exit Prism, it asks you whether you want to save the changes to every open file. If you click No, Prism
still saves the changed file in a special location for four days, and shows it in the list of unsaved files.
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This opens the Format Data Table window, which contains two tabs:
1. Table Format where you can make general changes to the data table format
In Prism tables, the rows and columns can have names or titles as they are called in Prism.
In most cases you do not use row titles so theres no point in adding or showing them. However,
when you want to label the data points on a graph, you have to add row titles because they are used
as labels for the data points.
2. Column Titles where you can change the names of the columns
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Another, simpler way to change the title of a column is to place your cursor in the appropriate cell
and just type a name:
Column titles are more important than row titles. You can skip row titles but you have to add column
titles to a table since they are used to:
Identify data sets in a data table when choosing analyses and viewing results
Label the X axis of graphs
Create legends for graphs
5.7.2. Change the name of a table
You may change the name of the entire table. Each time you create a new table, Prism will give it a
default name, e.g. Data1, Data2, Data3.... If you want to give a meaningful name to a table, right click
its default name in the Navigator window and select Rename sheet.
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Depending on the type of table you can select different items in the Sort drop down menu. Sort by X
Value is of course only available for XY tables. Sort by Row Title is only available for tables that
contain row titles. You can always reverse the order of the rows. Below you see the effect of
reversing the order of the rows.
Sorting of columns is not done in tables but you can do it in graphs so we will discuss it later in the
section on graphs.
5.7.6. Excluding data values
You can exclude data values from analyses and graphs:
Exclude (
Excluded values are shown in blue italics in the table so they stay visible:
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They are not included in analyses and they are not shown on graphs. From the point of view of
analyses and graphs, exclusion is the same as if you had deleted the value.
Note that you may only exclude data if you have a good reason for doing so!
Try Exercise 2A, 2B, 2C, 3, 4, 5, 6 and 7on the wiki page
Every analysis in Prism starts from the Create New Analysis window. To open this window:
Insert in Top menu -> New Analysis
Analyze (
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Selecting an analysis in this window opens a Parameters window that allows you to set the
parameters of the analysis. The Parameters window looks different for each analysis.
6.1. Transformations
In the Transform, Normalize section you can choose mathematical transformations (Transform,
Normalize) or table transformations (Prune rows, Transpose X and Y).
Transformations generate results tables that are colored in green and are stored in the Results
section of the Navigator:
You can't edit these tables, but the data sets on green data tables can be graphed and analyzed
further like any other table. Just click the Analyze button again. So you can first transform your data
and do a statistical test on the results.
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Prism contains a long list of predefined transformations, called standard functions but you can also
define a transformation function yourself by selecting User-defined X or Y functions.
For example standardization is done by selecting the built-in transformation: Y = Zscore(Y).
All mathematical transformations that are routinely used for transforming non-normally distributed
data into normally distributed data:
Log transformation: Y = Log(Y)
Square transformation: Y = Y squared
Square root transformation: Y = sqrt(Y)
Reciprocal transformation: Y = 1/Y
Many of the functions include the constant K. You can enter a value for K on the dialog (red). You can
enter one value of K for all data sets or a separate value of K for each data set. To enter different K
values for each data set, choose a data set, enter K, choose another data set, enter its K, and so on.
Rather than entering the value of K, you can hook a constant (see section 7).
If you entered replicate Y values, Prism can transform each replicate or the mean of the replicates.
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If you entered data as mean, SD (or SEM) and N, Prism will transform the error bars as well as the
mean if this is possible.
You can transform both X and Y values.
6.1.1.2. Transformations for pharmacology and biochemistry
Eadie-Hofstee, Hanes-Woolf and Lineweaver-Burk transforms are available for plotting enzymekinetic results, Scatchard transforms to visualize radioligand binding and Hill plots for dose-response
data.
You should use these transforms only as a way to display data, not as a first step in analyzing data.
For analysis use nonlinear regression on the actual untransformed data.
6.1.1.3. Normalization
Selecting the Normalize option allows you to set the parameters of the normalization:
You define the range (min and max) of the rescaled data: the new scale is set between 0% and 100%.
You can define 0% as the smallest value in each column, the values in the first row or to a value that
you specify. Similarly, you can define 100% as the largest value in each column, the values in the last
row, a value you specify, or the sum of all the values in each column. For replicates, 0% and 100% are
defined by the mean of the replicates. It is not possible to normalize each replicate separately.
You can choose to express the normalized values as fractions or percentages.
Note that X-values cannot be normalized: they are simply copied to the results table. Errors are
normalized appropriately.
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Analyses generate red results tables that are stored in the results section of the Navigator:
You can perform statistical analyses on these results tables allowing further analysis. Just click the
Analyze button again.
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Select Cumulative frequency distribution in the Create section. If you choose to tabulate the results
as fractions or percentages (in the Tabulate section), Prism offers you the choice of plotting an XY
graph with probability Y axis (under Graph type in the New Graph section).
You can only create QQ-plots for column tables. Since QQ-plots can only be created for the results
of an Analysis and not via a Graph, this restriction cannot be circumvented.
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The Shapiro-Wilk test does not work well when there are ties. When you have ties you should use
the D'Agostino-Pearson omnibus test. The Kolmogorov-Smirnov test is only included for compatibility
with older versions of Prism, and is not recommended. You can perform both the Shapiro-Wilk and
the Dagostino-Pearson test. Since both tests use a different method to check normality, they can
give different results!
Both tests return a p-value for H0: all values are sampled from a Gaussian distribution
p-value > 0.05: you have no good reason to believe that the data is not drawn from a
Gaussian distribution. So your data set passed the normality test.
p-value <= 0.05: the data are not sampled from a Gaussian distribution.
6.3.4. Comparing the mean or median to a hypothetical value
Prism offers a parametric t-test and a non-parametric Wilcoxon signed-rank test in the Inferences
section of the parameters of the Column Statistics. You can enter the hypothetical value to which
you wish to compare the mean (t-test) or median (Wilcoxon test). This value is often 0 or 100 (when
values are percentages) or 1 (when values are ratios).
Try Exercise 9 on the wiki page
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The Create and Tabulate sections allow you to choose to plot absolute (number of values), relative
(fractions or percentages) or cumulative frequencies.
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If you don't create a cumulative distribution, Prism gives you two choices of Graph type in the New
Graph section illustrated below: a histogram with points or a histogram with bars:
If you choose an unpaired test, the Parameters window does not change.
In the Choose test section you can choose to perform an F-test as part of the t-test. This might seem
strange since you have to specify if the variances are equal before Prism starts the t-test. The results
table of the t-test will then also contain the results of the F-test. If you perform a t-test on the
example data for the unpaired t-test, the results of the F-test are at the bottom of the results table:
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If the F test shows that the variances are not equal, its not a good idea to rerun the t-test but now
with Welch correction. Its better either to try to transform the data so that the variances are equal
or to use the Welch test routinely.
If you choose a paired test, you get the following Parameters:
You can choose between a paired t-test and a ratio t-test in the Choose test section of the
Parameters window. Clicking OK generates the following Results table on the example data for a two
sample t-test:
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Prism will test the effectiveness of pairing by calculating the Pearson correlation coefficient
between the control and treated group during the paired test. If the corresponding p-value is small,
the two groups are significantly correlated. This justifies the use of a paired test. If this p-value is
large, you should question whether it makes sense to use a paired test.
This is a bug in Prism!! The answer you see in the second column to the question Significant
correlation? (P>0.05) (in this example: Yes) is the answer to the question: Is p > 0.05? and not to the
question: Is the correlation significant ? So in the example above the p-value = 0.41 and thus p > 0.05
but this means that the correlation is not significant!
Try Exercise 11A on the wiki page
6.3.7. Multiple comparisons of two means in Prism
Prism allows you to perform one t-test per row of a grouped data table with replicates.
Select Multiple t tests - one per row in the Grouped analyses section of the Analyze Data window
Prism computes an unpaired t-test for each row and reports the corresponding p-value. There are
two ways to do this calculation:
Fewer assumptions: each row is analyzed individually, without assuming that data from
different rows are sampled from populations with identical variances. But as always when
doing a t-test, you do assume that in each row data from the two columns are sampled from
populations with the same variance. In this approach there are fewer degrees of freedom, so
less power, but you are making fewer assumptions.
More power. You assume that the data from both columns and rows are sampled from
populations with identical variances, so Prism computes one pooled variance. This gives you
more degrees of freedom and thus more power.
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Choosing the right option is not easy. If the data in different rows represent different quantities,
perhaps measured in different units, there is no reason to assume that the scatter is the same in all
rows. So choose "fewer assumptions". If they represent different conditions and the data are
measurements of the same variable, you might assume equal variances and choose "more power".
Prism corrects for multiple comparisons either by using a more stringent definition of statistical
significance (Bonferroni correction) or by controlling the FDR via the method of Benjamini and
Hochberg.
6.3.8. Comparing means of three or more groups
When the groups are defined by a single factor, each group forms a column in a column table and the
analysis is done by selecting One-way ANOVA (and non-parametric) in the Column analyses section.
Groups do not need to have the same size. Theres an example column table for one-way ANOVA.
This opens the Parameters window where you can specify the characteristics of your data sets.
Prism offers four ways to compare three or more data:
1. Unpaired data, normally distributed: One-way ANOVA
2. Unpaired data, not normally distributed: Kruskal-Wallis test
3. Paired data, normally distributed: Repeated measures ANOVA
4. Paired data, not normally distributed: Friedman test
Only the one-way ANOVA can handle data in mean, SD and N format, the other three tests cannot.
If you describe your data accurately in the Parameters window, Prism will pick the correct test.
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2. Multiple comparisons:
ANOVA will only tell you if there is a difference between the groups. It will not tell you which groups
differ. For this you need to do post tests in which you do pairwise comparisons of group means. You
can choose to do all pairwise comparisons or you can compare each group to one control group:
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So when the ANOVA reports a significant difference (p < 0.05) and you want to know which of the
groups are different, you have to choose a post test.
You choose a post test according to the comparisons you want to make:
All pairwise comparisons: Tukeys post test. Takes into account the scatter of all groups
resulting in more degrees of freedom and thus giving the test more power to detect
differences.
Compare all groups to one control group but not to each other: Dunnett's post test: Takes
into account the scatter of all groups resulting in more degrees of freedom and thus giving
the test more power to detect differences.
You select a set of groups to compare: Bonferroni or Sidak. Bonferroni is generally not
recommended because it is very conservative so you are more likely to miss real differences
especially when you have many groups. So don't use this test with more than five groups.
Both tests compute adjusted p-values and confidence intervals. If you dont need confidence
intervals use Holm-Sidak because it has more power than the other two tests. Note that you
should specify the pairs of interest before you collect the data. If you base your decision on
the data, it is not appropriate to use one of these tests for selected pairs. In that case, you
have to do all pairwise comparisons.
Fishers LSD test does NOT correct for multiple comparisons
Test for linear trend is used when the groups (columns) are arranged in natural order (e.g.
increasing dose) and you want to test if there is a trend e.g. values increase when you go
from left to right column.
3. The Options tab allows you to specify the method for multiple comparisons correction and the
content of the results that are generated by this analysis.
The tests that you can choose here depend on the choices that you have made in the Multiple
Comparisons tab. If you choose to compare every mean to a control mean, you can make the choices
listed above. If you choose to compare every mean with every other mean, you'll be choosing from
the list that is shown below.
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One-way ANOVA
The analysis will generate two results tables:
1. The results table called ANOVA:
The P value tells you if H0 (the groups all have the same mean) is true. In this example, there are at
least two groups that are different (P < 0.05).
In the ANOVA Table:
SS of Treatment (between columns) represent the variation between groups
SS of Residual (within columns) represent the variation within groups
Each of these two measures is divides by its corresponding degrees of freedom (df) to generate the
MS you see in the table:
MS = SS/df
When you calculate the variance between groups:
When you calculate the variance within groups:
df = number of groups - 1
df = number of subjects - 1
If H0 is true, there is no difference between the groups and the variation between groups should be
more or less equal to the variation within groups so F = 1. So the larger F, the more the groups differ.
R square is the fraction of the total variance that is attributable to differences among group means. A
large R square means that most of the variation is caused by the treatment that defines the groups.
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If each group has at least five values, Prism will test if the groups have equal variances using
Bartletts test and the Brown-Forsythe test. H0 states that they have equal variances.
2. The results table called multiple comparisons shows which groups differ
You see that the adjusted p-value is not reported because we havent explicitly asked to report it,
instead the test reports 95% confidence intervals for the differences between the groups. If you
want to know the adjusted p-values you can select this option in the Options tab:
When you select this option, the adjusted p values will be reported.
When the columns are ordered, e.g. increasing dose, you can test if the column means increase or
decrease systematically as you go from left to right column by selecting the Test for linear trend in
the Multiple comparisons tab:
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This test performs a linear regression between column mean and column number. H0 states that
there is no linear trend. Prism reports the slope and the regression coefficient (R squared) of the
regression as well as the p-value for the linear trend:
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The results table of the repeated measures ANOVA is very similar to the one generated for regular
one-way ANOVA.
Just as in the paired t-test, Prism tests if the matching was effective. For this test, H0 states that the
matching is not effective. So if the p-value is high you could consider an ordinary one-way ANOVA.
Another difference is that there are three sources of variability: between columns (groups), between
rows (individuals) and random.
For repeated measures ANOVA, you can use the same post tests as in regular one-way ANOVA.
Prism performs type I ANOVA, which basically means that you can only draw conclusions for the
groups that you have collected data from. Prism cannot perform type II ANOVA, in which you assume
that you have studied two or more groups from an infinite number of possible groups and you want
to draw conclusions that are valid for ALL the possible groups.
Kruskal-Wallis and Friedman test
Following the nonparametric Kruskal-Wallis or Friedman test, Prism can perform Dunn's post test for
comparing either all or selected pairs of groups. Although non-parametric, these tests were
developed for continuous data, thus expecting that each data value is unique. If you have ties in your
data, this expectation is not met and the test cannot calculate an exact p-value. Prism will use an
approximate method to calculate an approximate p-value.
6.3.9. Correlations
Prism calculates correlations for XY or Column tables. Remember: in an XY table you may not
calculate the correlation between X, a variable that you have manipulated, and Y, a variable that you
measured. You may only calculate correlations between two variables that you measured. However,
Prism doesnt care about this and allows you to calculate correlations between X and Y columns.
To calculate correlations, open the Analyze data window and select correlation:
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If you have more than 2 columns, you can specify between which pairs of columns you want to
compute the correlation. You can choose between Pearson and Spearman correlation depending on
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the distribution of your data. Prism also computes a P value for the correlation. If the p-value is small,
you can reject the hypothesis that the correlation is due to random sampling.
Try Exercise 12 on the wiki page
7. Information sheets
Info sheets are used to record experimental details and constants that you used in analyses. These
constants can be used (hooked) in analyses.
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You can simply type info in the information sheet. For instance, suppose you want to use a certain
constant for transformation of your data, just go to an empty row and type transformation constant
at the left and its value at the right.
When you click the hook icon, the Hook constant window opens:
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You can select a constant from a list of the constants that you entered in the info sheet.
To insert an Info constant into a graph title, legend, or text object:
Click the info (
8. Graphics
By default, Prism automatically creates one graph for each table you create, but you can plot
multiple graphs for a table. And you can plot data from multiple tables on one graph.
For each table type Prism displays all the choices on a row that you can horizontally scroll through.
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Although Prism seems to imply that you can only create certain plots for certain table types, this is in
fact not the case. You can create all kinds of graphs for any table type.
You can specify the statistics that are plotted on your graph in the Plot dropdown menus:
When you select a graph type, its name is displayed below (green rectangle).
For XY tables with one Y column without replicates, the scatter plot is straight forward but you can
also generate scatter plots for XY tables with multiple Y columns as in the example below.
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If you do have replicates, you can choose to plot the mean and error bars instead of the individual
data points. For the error bars you can choose between SD, SEM and 95% confidence intervals.
But you can also choose to plot individual data points: either Aligned or Staggered.
Aligned means that data points with the same X and Y values will be plotted on top of each other
whereas Staggered will plot them next to each other.
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You cannot switch the X and Y-axis to generate a horizontal plot. For this you have to select Grouped
in the Graph family and choose the horizontal dot plot.
You have to plot the mean and error, Prism doesnt allow you to plot the individual data points.
If you want to plot individual points for grouped tables, you can select a scatter plot in the Plot
individual values tab:
In the figure on the previous page the two groups, control and treated, are separated but you can
also group the data according to the second grouping variable.
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At the left is an interleaved scatter plot, where the groups are plotted next to each other. You can
also superimpose the groups (put them on top of each other) as you can see at the right.
For column tables, the scatter plot can show every data point along with lines for mean and error
bars for each column but you can choose to leave out lines and error bars, which is often more clear.
Below is an example of a scatter plot for two columns showing individual data points AND the mean
(colored line) AND the error bars (black bars), generated in Prism.
You can also choose a horizontal dot plot displaying the mean and error, which is also very clear.
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This is the only bar chart in Prism that will place the bars according to a coordinate. In bar charts for
all other tables the X values are considered as groups labels and plotted with equal spacing.
For column tables, each column becomes a bar with equal spacing between the bars:
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If descriptive statistics are available you can specify which statistics you want to plot.
For grouped tables each column represents a different type of bar. In the example below there are
three columns giving rise to three types of bars: cats, dogs and hamsters. Rows can be plotted along
the X or along the Y axis. Each row is a different instance of that bar, e.g. in the example below there
are four rows. For each row, three bars are plotted one for cats, one for dogs and one for hamsters.
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A contingency table is exactly like a grouped table except that it doesn't have sub-columns for
replicate values so the bar chart will be similar to that of the grouped table except that it will lack
error bars.
8.2.4. Box plots
Prism links box plots to Column and Grouped tables.
Box plots are based on the following descriptive statistics: min, Q1, median, Q3, and max. These
statistics are also called Tukeys five number summary, so this is what the Tukey in the Plot
dropdown menu points to. So the Tukey option creates a boxplot with whiskers at 1.5 * IQR from the
box and outliers.
The Min to Max option creates a box plot with whiskers at the minimum and maximum of the data
set. Normally, you would prefer Tukey over Min to Max but if a data set is very small its a good idea
to choose Min to Max and show all the data points on the plot.
You can change the order of the boxes by selecting a separate box plot instead of an interleaved one.
Try Exercise 14 on the wiki page
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Format Graph (
) in Change section of upper toolbar
Double-click any aspecific point on the graph (not a data point, axis, title)
Right click a specific part of the graph e.g. axis, legend, data point
Change in Top menu -> feature that you want to change
The Format Graph window contains three tabs, the two important ones are:
Appearance allows you to specify what the graph should look like
Data Sets on graph allows you to specify the data shown on the graph
The Appearance tab contains the following sections:
Data Set determines to which data points in the graph, the changes will apply. You can choose to
change features for all data points or for specific subsets.
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Appearance allows plotting different statistics or none. You can choose to show individual replicates
instead of error bars by selecting Each Replicate. To show error bars select Mean/Median and Error.
If you select to choose error bars: you can specify in the Plot dropdown menu which statistic you
want to use for the error bars (standard deviation, standard error of the mean, confidence intervals).
In the Show symbols section you can specify symbol type, size, and color.
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If you pick an open symbol, you have to select a border thickness. Check the Clear option if you want
overlapping points to show through the open symbol. Use the Shape drop-down menu to select a
symbol. Use the More option to use any character from any font on your computer as a symbol.
Use Show bars/spikes/droplines if you want to drop vertical bars from your data points. The bar will
start at the X-axis or whatever Y value you choose and go up (or down) to the Y position of the point.
This is useful when you want to create a bar graph where each bars position is determined by its X
value. It is also useful when you want to combine points and bars on one graph as shown below.
In the Show error bars section you can specify the appearance of the error bars. Choose dotted or
solid lines to show an error envelope as in the graph on the left. Use the Show Area Fill button to add
fill between the limits of the error envelope as in the graph on the right.
In the Show connecting line/curve section you can choose the line's color, style, thickness, and
pattern. This section includes choices to have your connecting start either at the first data point or
the graph origin, whether to leave gaps in the connecting line next to the symbols, and whether to
plot lines connecting each replicate for repeated measures in corresponding data sets. Don't confuse
the connecting line which always goes from data point to data point with a best-fit curve or line
created by regression. If you have fit a curve through the data, Prism created the curve as a separate
data set. Add the data set to the plot.
The interface of the Format Graph window is different for different graph types and for the same
graph type on different table types. The example shown above is for scatter plots of an XY table,
whereas the Appearance section of a scatter plot of a column table looks like this:
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Depending on the graph type, you can change different aspects of the graph, e.g. for graphs with
boxes you can change the appearance of the boxes, for graphs with dots you can change the
appearance of the symbols. If there are error bars on the graph, you can change their appearance
too. We will just show the example of the Bars section. The other sections are very similar.
In Bars or Bars and boxes (depending on the graph type) you can specify the color, border, pattern
and line width of the border of the bars/boxes:
The additional options section allows you to remove the legend and to label each data point on the
graph with the title of its row (provided you have added row titles of course).
You may add columns from other data sets to the graph.
If you click an individual data point, you can change a feature of that individual symbol or bar:
-> Format this point -> Feature that you want to change
But you can also change a feature of the complete column that data point belongs to:
-> Format Entire Data Set -> Feature that you want to change
Or you can a feature of the entire table:
-> Format All Data Sets -> Feature that you want to change
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This window has five tabs. On the Frames and Origin tab you can change the Height and the Width of
the Axes.
The X-axis tab allows you to specify the appearance of the X-axis. In an XY-table the X values you
enter into the X column are plotted on the X-axis. The X axis, in this case, is numerical. If the X values
are not evenly spaced, then the points won't be equally spaced. In the X-axis tab you can change the
range, appearance and location of the ticks. To change the range of the X-axis: uncheck the
"Automatically determine the range and interval" box and enter the minimum and maximum value
that you want to plot on the axis. You can also hook a range limit to an info constant.
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For Column tables, column titles are used as labels on the X-axis. For grouped tables, row titles are
used as labels on the X-axis. In these two cases X values are not numerical and they will be equally
spaced on the axis. For these table types, you can only change the appearance of the ticks in the X
axis tab.
The Y-axes tabs are very similar to the X-axis tab. Since Y-axes are always numerical you can change
the range, appearance and location of the ticks for all table types.
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Another useful feature of this window is on the Titles & Fonts tab, where you can choose to show or
hide the titles of the graph and the axes and to change the direction of the title of the Y-axis.
Set this to horizontal since horizontal text is much easier to read than vertical text.
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Prism will give you the option of deleting the data and add your new data. You can clone from any
project that is open, used recently, or from projects that you saved as examples.
http://www.graphpad.com/guides/prism/6/user-guide/clone_a_graph.htm
Then you have to choose the graph that you want to use as a template
-> Next
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Select the properties of the template graph you want to apply to your current graph -> OK
9. Repeating analyses
We have already seen two strategies to easily repeat an analysis on new data (cloning a graph,
duplicating a family) but there is still another strategy that you can use. Suppose you've already
entered data onto an empty data table, it is too late to clone and too late to duplicate a family. Prism
provides a great tool to solve this problem: the Wand (Wizard to ANalyze Data). Use it to analyze
(and graph) the new table just like you already analyzed (and graphed) an existing table, see:
http://www.graphpad.com/guides/prism/6/user-guide/use_the_wand_to_repeat_analyse.htm
Use
to export tables in various formats. Works for data tables, result tables and graphs.
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