Frequently Asked Question for PLS Program ========================================= Q: Besides GUI interface, how can I get the meaningful information directly PLSgui result file (i.e. *result.mat)? A: Yes, you can. Since some variables are only used internally by the program, and some other variables do not have a meaningful name, I am creating a mapping list here for your convenience. First, you use "load" command to load the result file into MATLAB command window, then copy the block below and paste it to the MATLAB command window: %%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%% clc %%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%% num_of_group = length(SessionProfiles) SessionProfiles{1} % listed all subjects with path in 1st group num_of_permutation = perm_result.num_perm num_of_bootstrap = boot_result.num_boot switch method case 1 type_of_analysis = 'Mean-Centering PLS' case 2 type_of_analysis = 'Non-Rotated Task PLS' case 3 type_of_analysis = 'Regular Behavior PLS' case 4 type_of_analysis = 'Multiblock PLS' case 5 type_of_analysis = 'Non-Rotated Behavior PLS' end confidence_level = boot_result.Clim condition_name = cond_name selected_condition = find(cond_selection) %%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%%% Q: Can I enter decimal number in the "Onsets" field for MRI study? A: Yes, you can. However, since the smallest unit is a scan (i.e. 1 TR), the program will round your decimal number to integer. If you selected "Use HRF instead of Block length", we adopted some feature from SPM. Instead of only using 1 time bin, SPM increases the Microtime resolution 16 times. This may bring some benefit if you input decimal onset time directly, although eventually the length of convolved onsets is still the number of scans (i.e. one scan is still the smallest unit). However, if you input decimal onset time directly, please make sure that your slices must be in sequential order, instead of in interleaved order. Also, please make sure that your slices must be in ascending order (i.e. 1st slice is at bottom) instead of descending order or rounding order. Otherwise, you must enter integer for the onset time. For other kinds of fMRI in PLSgui, since they are either based on temporal window or block length, there is no chance to take decimal onset time. If you do input decimal onsets, they will be rounded to the bottom slice using "round" function (before Feb 24, 2011, they were rounded using "floor" function). Q: How many LVs do I have? A: The number of LVs that generated by our PLS software is determined by which option that you choose to run PLS: For Mean-Centering PLS: nLVs = nConditions x nGroups; For Non-Rotated Task PLS: nLVs = nContrasts; For Regular Behavior PLS: nLVs = nConditions x nGroups x nBehaviorMeasure; For Non-Rotated Behavior PLS: nLVs = nContrasts; For Multiblock PLS: nLVs = nConditions x nGroups + nConditions x nGroups x nBehaviorMeasure; There are also several restrictions: 1. Since nConditions only provide nConditions-1 degrees of freedom, the maximum number of contrasts is limited by nConditions; 2. In order to run Behavior PLS, you need to have at least 3 subjects; 3. In order to run Mean-Centering PLS, you need to have at least 2 conditions. However, if you have only 1 condition and you have more than 1 groups, our PLS will reconstruct datamat so that it will become single group with multiple conditions. Q: Which side is the left side of my image? A: If your image data is in NIfTI format, PLSgui always load, save, and display your image in RAS orientation. i.e. Left to Right on X axes, from posterior to Anterior on Y axes, and from Inferior to Superior on Z axes. Therefore, left side is the left side of your image. However, if your image data is in ANALYZE format, please refer to the answer below to manually adjust the orientation of your image. Q: Can I use Analyze image with different orientation? A: Yes, a new feature has been added to the version of Octobor 29,2004. This allows people to use Analyze images in any orientation for PET, E.R.fMRI and Blocked fMRI. In "Create Datamat" window, a new button named "Check image orientation" is added beside the "Create" button. Once you click this button, the 3-View of one of the subject images will be displayed in the "Change Image Orientation" window, and you can re-orient it by click the "Re-orient" button. Once you click the "Re-orient" button, it will ask you to input the orientation of the CURRENT DISPLAYED image. If you did not get the correct orientation, you can do as many times as you want to make it standard Analyze orientation, i.e. from Left to Right in X orientation, from Back to Front in Y, and from Bottom to Top in Z. If you closed the "Change Image Orientation" window, you can re-open it by click the "Check image orientation" button in "Create Datamat" window again. Q: Can I use 4D NIfTI or Analyze image data? A: Yes. For E.R.fMRI or Blocked fMRI modules, you can use 4D data directly. For other modules, all you have to do is to run a small program called "expand_nii_scan.m" under command promot, and expand all the 3D scans into different folders. Q: Does PLSgui support ANT average binary data? A: Yes. The "load_ant.m" program in PLSgui is based on openlib library "cntopenlib.zip" and additional information "avr.txt" file that are released by ANT technical support. Since January 2004, ANT's average data file has been changed, and the history section is included in its header. This info is not included in the "cntopenlib.zip"; however, it is in the "avr.txt" file released by ANT's support people. Some people used "avr2asc" provided by ANT's EEProbe_3.3.120 to convert ANT's average data to plain text file. However, the disadvantage is that the "avr2asc" only supports new version of ANT's average data on Linux and Mac platforms, and does not work under MATLAB. Other programs relied on their "cntopenlib.zip" (e.g. Robert Oostenveld's "read_eep_avr" that is used by EEGLAB) can only support old version, and they are compiled in mex or dll file. You must first use "avrstrip" in EEProbe_3.3.120 to convert your new version of ANT's average data to old version before you can use "read_eep_avr" to load them. The "load_ant.m" is the only explicit .m program so far (April, 2007) that supports both old & new versions of ANT's average data file on any platform. Q: In my MEG data, the number of channels and their corresponding names do not match up with any of the pre-specified system. How can I read in my data when channel configuration and names do not match? A: Up to this version (5.0704231) We only have 4 scalp electrode location systems in PLSgui. They are the ones that we are using most frequently. There is a way to add any system by yourself, as long as you know the electrode names with their corresponding x and y locations. Since PLSgui will normalize your x and y location to Xmin/Xmax and Ymin/Ymax, you don't need to worry about the exact locations, and the relative ones will work. Here is the details of how to add a Standard 10-20 EEG System with 19 cap electrodes: 1. First step: Put all electrode names on a piece of grid paper, and make sure that they are spatially located appropriately. 2. Second step: Select an origin for XY coordinates. You can pick any point (on or off any electrode) as your origin. For example, Cz is a good selection, the most lower left grid is also a good selection. 3. Third step: Use a ruler (or count the grid) to measure the x and y location from the origin. 4. Forth step: Pick any of the 4 PLSgui electrode systems for your system. In PLSgui, each system is determined by a "erp_loc*.mat" MATLAB file. So we have: "erp_loc_besa148.mat", "erp_loc_egi128.mat", "erp_loc_egi256.mat", and "erp_loc_ctf150.mat". The default system for "Edit Channel Order" window is: "erp_loc_besa148.mat". So, it will be convenient for you if you pick this one. The whole idea to use your own system is to create an electrode systems MATLAB file with the same file name as the one used by PLSgui. Let's assume you picked "erp_loc_besa148.mat", once you save your own "erp_loc_besa148.mat" in the same folder as the one you will save session/datamat/result file, PLSgui will load your own "erp_loc_besa148.mat", instead of the one from PLSgui program folder. All 4 PLSgui electrode system MATLAB file in PLSgui contain two variables, one is char array "chan_nam", the number of rows is the number of channels and the number of columns is the longest length of the electrode names; the other one is double array "chan_loc", the number of rows is the number of channels and it only has two columns. Column 1 is for x values, and column 2 is for y values. Now continue with this example: chan_nam=[ 'Fp1' 'Fp2' 'F7 ' 'F3 ' 'Fz ' 'F4 ' 'F8 ' 'T7 ' 'C3 ' 'Cz ' 'C4 ' 'T8 ' 'P7 ' 'P3 ' 'Pz ' 'P4 ' 'P8 ' 'O1 ' 'O2 '] chan_loc=[ -3 10 3 10 -8 6 -5 7 0 8 5 7 8 6 -10 0 -7 0 0 0 7 0 10 0 -8 -6 -5 -7 0 -8 5 -7 8 -6 -3 -10 3 -10] % Assume that your current directory is the one that % you will save session/datamat/result file, then: % save erp_loc_besa148 chan_loc chan_nam Now you have your own system in "Edit Channel Order" window if you are under "ERP/BESAThetaPhi" system, and you can also pick channels that is used in the ERP data. Q: I am not using MATLAB in MS Windows platform now. When I click "How to use this window?" under help menu, I only get "To learn how to configure your Web browser type 'help docopt'" message in MATLAB command window. When I did so using MATLAB in MS Windows platform, I can see that the help is nicely opened in my default web browser. Does it mean that this menu can only be used in MATLAB in MS Windows? A: This menu should work on both Windows and Unix (Linux). I have not tested it on other OS. However, you could still encounter error (the message in your question is automatically displayed by MATLAB) because of the following reasons: 1. You are using MATLAB version 5. It does not have a MATLAB desktop, and it uses the current terminal for commands. 2. You specified -nodesktop or -nojvm when you launch MATLAB. 3. You do not have any web browser in your system, or you did not specified properly in "docopt.m" file. In latter case, please type 'help docopt'. In any case, in fact, I recommand PLS user to manually open your favorate web browser, and view our PLSgui User's Guide from PLS web site: http://www.rotman-baycrest.on.ca/pls/UserGuide.htm Q: Why the figures in UserGuide.htm can not be displayed? A: Because those figures took hundreds of kilobytes of the space, I have to leave them in our website. As long as your computer have internet access, you will be able to see them properly, even if you browse "UserGuide.htm" that is on your local computer. Q: I wrote a batch file to create fMRI datamat. I have 5 conditions and 2 runs. Since I set "across_run" keyword to "0" (merging data within each run), I will have 10 conditions. I listed all 10 after 10 "cond_name" keywords. When I run the batch, I got error message saying "Index exceeds matrix dimensions". A: You can only list your 5 conditions after 5 "cond_name" keywords. This is because the program will expand your 5 conditions in your 2 runs to something like Run1Cond1, Run1Cond2, Run2Cond1, etc. Q: I preprocessed my data in FSL, where left is R and right is L. I then entered the normalised, slicetime- and motioncorrected data into a PLS analysis. When we now look at the results, where is left and right? As in FSL? A: Let's use command: nii = load_nii('filename.nii'), where "filename.nii" is the one from FSL: 1. If you find nii.filetype is 0, it means that the image orientation is as is in the data. 2. If filetype is not 0, and there is no "rot_orient" & "flip_orient" field under "nii.hdr.hist", it means that your data is already in RAS, and my NIFTI package did not convert your data. 3. If file type is not 0, and there is "nii.hdr.hist.rot_orient" or "nii.hdr.hist.flip_orient", it means that your data has been converted to RAS (Left is L) by NIFTI package. Q: What should the contents of the input file for "multiple voxel extraction"? I created a text file with 1 XYZ coordinate per row. However, I keep getting an error message. A: That's the correct contents. However, please make sure that the XYZ coordinates should be in unit of millimeter. (Note, we did use absolute voxel location before 15-SEP-2006. You can use script "xyz2xyzmm.m" to easily convert absolute voxel location to voxel location with unit of millimeter.) Q: When I use the multiple voxel extraction, does it output the baseline-adjusted values or the raw image values? A: It outputs the baseline-adjusted value (from *datamat.mat), and there is no raw image values kept. If you need raw image values, you have to load images by yourself, and extract only the voxels indexed by the XYZmm file you provided. Q: I would like to get intensity values at particular coordinates from the raw image. How do I load image and extract the voxel intensity from XYZmm file? A: Let's say your raw image is: filename = 'SCAN.img'; or filename = 'SCAN.nii'; and you would like to get the intensity values at particular coordinates from it. First, you need to load the image by yourself: A = load_nii(filename); Then, display the image: view_nii(A); Since you have already prepared XYZ(mm) in voxel file, you need to switch the selection of "Axes Unit" from "Voxel" to "Millimeter", and then enter each XYZmm value in "[X Y Z] at crosshair" field and click TAB key. The value displayed in "Value at crosshair" is the intensity value at the particular coordinate you entered. If you have many coordinates, you can use the simple script below to automatically extract intensity values list from locations list in your XYZmm file: intensity=[]; xyz = xyzmm2xyz(xyzmm, result_file); for i=1:size(xyz,1) intensity=[intensity;A.img(xyz(i,1),xyz(i,2),xyz(i,3)]; end Q: In E.R.fMRI and Blocked fMRI datamat create window, what does the "Normalize data with ref.scans" check box mean? A: This check box is checked by default to remove the low frequency noises and scale the data. The formula for this check box is very simple: dataset = (dataset - ref_dataset) * 100 ./ ref_dataset In this way, activity for each event is expressed as a %change from the reference scan (i.e., baseline) similar to what is done for ERP, where activity is expressed as a change from prestimulus baseline. Please keep this check box checked unless you have a good reason not to do so. Q: Also in E.R.fMRI and Blocked fMRI datamat create window, when the "Single Subject Analysis" check box was checked, the red words "Number of onset must be same" were presented in the bottom of this window, however,when this check box was not checked, session session file could be created successfully. Why? A: For single subject analysis, we actually treat each onset as a "subject" (in the sense of our "subject-in-condition-in-group" format). This is why we need you to keep onset number same, because the number of "subject" should be the same for each "condition" in each group. If you uncheck the single subject analysis, all the onsets for each condition will be averaged together, so there is no need to keep onset number same. Q: I have three conditions (positive, negative and neutral) and I want to compare the effect of positive vs neutral, if the "Mean-Centering PLS" is cheked, how do I "deselect" conditions in the window of "PLS Analysis"? A: Click "Deselect Conditions" under "Deselect" menu, and deselect the "nagative" condition. Q: What does the "Non-Rotated Task PLS" check box in "PLS Analysis" window means? A: In "Non-Rotated Task PLS", we directly used crossblock between the contrast data and the task mean of your fMRI data as brainlv, and directly used sum of squre of the crossblock for singular value. There is no PCA computation involved, and there is no need to do procrustean rotation for the result. This is why we call it "Non-Rotated Task PLS". Q: If the "Non-Rotated Task PLS" check box is checked, how do I design the contrast? A: Click "Contrast" menu in PLS analysis window. Click "Using Helmert Matrix" under the "File" menu of the contrast window. The freedom of contrast is number of conditions minus 1. For example, if you have 3 conditions, you can have 2 contrast. You can either take the Helmert Matrix value in the text box, or re-enter your own contrast value (e.g. [-1 1 2], [1 -2 3] etc.). At the end, you have to go to "File" menu to save your contrast data into a file, and this file name will be entered into the "Contrast Data File" in the PLS analysis window (you can only enter file name under "Non-Rotated Task PLS" option. Q: I have completed a group analysis with seven subjects under three conditions(positive, negtive and neutral) using Helmert contrast, when showing the result, how could I find the the values of latent vectors for design contrast? A: Click "Design Scores and LVs Plot" in the "Windows" menu of the result display window. The Design Scores for each latent variable reflect the impact of your contrast data. Q: In my result the "Brain LV" view is very different from the "Bootstrap Ratio" view with default threshold, and which view do I prefer to support my experiment result? A: The "Bootstrap Ratio" view gives you a "stability map", so you can focus on the high positve (or negative) voxels in the "Bootstrap Ratio" view. However, "Brain LV" view is the one to support your experiment result. The positive (negative) value should be consistent with the one in Design Scores plot. Q: I have more than one runs in my MRI study. However, for some runs, I do not have all the conditions. How should I fill out the onset field? A: If you want to exclude certain conditions for certain runs, you can simply put a -1 for the onset field, (and put a 1 for the length of Blocked fMRI). Then choose "Merge data of each condition: across all runs" to create a datamat. Please ignore the command line message saying "Scans -01 not included due to out of bound". You can not choose "Within each run only", since you do not have all the conditions filled for every runs. In additon, for each condition, you must have at least one valid onset (not -1) in one of your runs. Q: Why the results I obtained using Matlab 7 can not be loaded under earlier version of Matlabs? A: In order to load results generated using Matlab 7 on earlier versions of Matlab, a special Matlab 7 switch must be included during the saving process. Since such a switch is not coded in the package, a small program called 'matlab7to5.m' is included to covert the .mat files generated from Matlab 7 to be loaded on earlier versions of Matlab that are above Matlab 5. Q: I got "divide by zero" error when I tried to run Behavior PLS analysis, while there is no error when I ran task PLS. A: There are two reasons that could bring you into this trouble: 1. You must have at least 3 subjects to do the behavior PLS. 2. Values should not be all the same for behavior data. Even if they are really all the same, you must add (or minus) a slight difference (e.g. 1/1000) to make it work. Q: Is there any restriction on file name? A: Yes, but not many. There are 3 kinds of major files that will be used in this program. They are session file, datamat file, and result file. The 3 major files must be in the forms of: *_MODULEsession.mat *_MODULEdatamat.mat *_MODULEresult.mat You can replace MODULE with any of PET, ERP, fMRI, or BfMRI, depending what PLS are you working on, and the dialogbox will always prompt you with the correct MODULE. You can not insert your own string into the above forms and break it. However, you can always use your own string before "_MODULE", which is called "prefix" in the program. I do not recommend you to use any blank space or symbol characters like "+ - * / % ^ $ ' ..." etc. inside the string. In addition, you should not rename the filename manually by yourself, since this will break the filename linkage stored inside the above major files. Q: When I opened the result file, it crashed and told me that the datamat file was unable to open. (No, the datamat file was not in the current directory.) A: In the early version of the PLSgui program, we used the stored 'pls_data_path' variable to locate the files. However, this also caused trouble when people want to copy those datamat files from directory to directory, and even from one machine to the other. So, we decided to use the "current directory" rather than the stored 'pls_data_path' variable for "Run PLS" window and "PLS result" window. If you are not starting the program from the "current directory" where those datamat files sit, you can always change to "current directory" in the very first window by clicking "Change Current Working Directory" under "Tools" menu. Q: When I add or edit subject directory for ERP and PET, I saw an edit box called "Number of character for subject initial". What should I enter? A: For ERP and PET data, each subject has its own directory. The data files inside the subject directory stand for different conditions for this subject. The name of these files can be arbitrary; however, if you give "consistent naming across subjects", it will bring you convenience to match the conditions. Here is what the "consistent naming across subjects" means: 1. All subject files consist two portions, subject initial portion and condition name portion. e.g. SubjInit1_CondName1; 2. Across all subject directories, the condition name portion should be the same for the same condition; 3. Within each subject directory, the subject initial portion should be the same for all conditions of this subject; Here is how to make this feature work for you: 1. Enter the length of the subject initials in the "Number of Characters for subject initial" box. You can disable this feature by leaving this value as -1. 2. Click Add or Edit button in the same window (Edit Subject Directory window), and a new window called Subject Directory Detail will open. 3. Select one of subjects that will be used in this datamat group (Note: Remember that only select 1 subject this time). 4. If you have "consistent naming across subjects", you will notice that "File names are the same across subjects" check box is checked. Uncheck it to disable this feature. 5. Go to right hand side, select correct subject files that match the inputted conditions at their left side. Make sure that no subject file can be duplicated. 6. If you have multiple subjects in this datamat group, you can now select the rest of subjects by holding Shift or Ctrl key combination while selecting (Note: You can not do so before this step). 7. Click Done button when you finish, and you return to "Session Information" window. Q: I received a "Subject file name convention is not consistent" message when I try to edit a subject. However, all my subject file name convention are consistent. Is this a bug? A: No, it is not a bug. When you edit a subject with "File name are the same across subjects" checkbox selected, the program expects that you are trying to use a different subject. You should manually do the mapping between subject files and input conditions by yourself. If the mapping you selected does not follow the other subjects you selected, the program will display such an error message. If you definitely need such a mapping, then the "consistent naming across subjects" rule can not be kept, and you will have to uncheck the "File name are the same across subjects" checkbox. Q: All my subject files have consistent naming convention, and I also input the correct length in the "Number of Characters for subject initial" box. Why the "File names are the same across subjects" checkbox is grayed? A: Please check the following 2 possibilities: 1. All files under any subject directory should follow the "consistent naming across subjects" rule. If one of them does not follow this rule (e.g. WS_FTP.LOG), the "File names are the same across subjects" checkbox will be grayed. 2. Don't let external program affects your subject initial. For example, there is a subject file with subject initial "S1" with length of 2 characters. When you normalize this file, it may become nS1 with length of 3 characters. You will have to rename the file name if this happened. Q: Does the length of the data file name matter? A: No. It can be any length. However, please make sure that subject initial portion should be the same length if you would like to use the "consistent naming across subjects" feature. Q: Does the letter case of the data file name matter? A: No. It is particularly designed for case insensitive. So you can call the first data file name "JSdata1", and the second "jsData2". Q: What can this PLS Program do? A: PLS indicates Partial Least Squares (PLS) analysis method. It is based on the assumption that the focus of analysis is on which aspects of the signal in one matrix are related directly to signals in another matrix. This PLS program is applied to neuroimaging data, such as PET, ERP, Event Related fMRI and Blocked fMRI. It has been used to identify the task-dependent changes in activity, changes in the relations between brain and behaviour, and to examine functional connectivity of one or more brain regions. The similarities and differences in brain-behavior relations can be identified from the resultant patterns. Q: What data should I prepare for analysis? A: The data can be obtained from PET, ERP, or fMRI scan images. For PET and ERP data, you put different condition of data for each subject into its corresponding directory. For fMRI data, you put each run of data into its corresponding directory. Q: I made some changes, but it disappeared. Why? A: Any changes you made on the edit box need to be recorded. This can be done by clicking "Enter" key on your keyboard, or simply click anywhere else on the window to let the focus switch off from this edit box. If you did not let the focus switch off from this edit box, the change will not be recorded. Q: What should I enter for "Edit Behavior Data"? A: For PET and ERP data, the number of rows for behavior data should be the number of conditions times the number of subjects, in the order of subjects in conditions: cond1/subj1 cond1/subj2 ... cond2/subj1 cond2/subj2 ... For fMRI data, the number of rows for behavior data should be either just the number of conditions, or the number of conditions times the number of runs, depending on whether you choose of "across run" or "within run" when you create datamat. The order should be either all conditions, or runs in conditions: for across run: cond1 cond2 ... for within run: Run1Cond1 Run1Cond2 ... Run2cond1 Run2Cond2 ... The column of the behavior data should correspond to the different sets of the behavior measures. Q: When I exported data for behav analysis for PET module, I saw the exported file name looks like "?_PET_grp?_behavdata.txt". For the both E.R.fMRI and Blocked fMRI modules, however, it looks like "_fMRI_grp?_subj?_behavdata.txt". Why the PET module does not include the subject information? A: For PET module, each datamat (and session file) represents a group that already contains subjects. However, for fMRI, each datamat (and session file) represents a subject. So, for fMRI, we need behavior measure of each subject in the group in order to add it into datamat (from session profile). For PET, we need behavior measure of each group. Q: Can I rename the session / datamat / result files manually? Why the corresponded session file also changed its name, after I save as fMRI datamat into another file name? A: We do not recommend you to change name manually. For fMRI, there is some internal processing between session / datamat files. That's why the session file name will be changed after you saved datamat file into another name. Q: After deselect the conditions, why the behavior data I loaded before running analysis get lost (or still use the one from datamat)? A: Because the rows of behavior data is determined by the conditions you selected. So, you need to load behavior data after you deselect the conditions. Q: When I generate fMRI datamat, I got something like this: Scans 147 for the condition 3 of run1 are not included due to out of bound. Do you have any idea what this means? A: When you enter onset data, make sure they are in the range. In most case, you encounter this problem because the onset data you input exceeds the maximum range. The minmum value is started from 0. However, The maximum value you can enter is: Number_of_Scans - Temporal_Window_Size_if_fMRI - 1. E.g.: If you have 180 files, and window size is 8, the maximum value in onset data is 171. Q: What is the lag number? Is there anywhere I can enter lag values? A: In E.R.fMRI, lag number is the index of the temporal window, and it starts from 0. You decide the temporal window size in "Generate ST Datamat" window. The unit of temporal window size is scan, and its default value is 8. In "LagXYZ" field of result window, you can enter lag value, as well as XYZ coordinates in voxel. Q: In our lab we use AFNI in the processing of fMRI data but I would like to use the PLS program to analyse an event-related dataset and further progress to conducting a functional connectivity analysis. My problem is that the PLSgui seems to need separate image files for every time volume (a separate image file for every 'scan' in a run), however, using AFNI, the raw 2D timeseries is transformed into a 3D dataset, upon which the motion correction, edge-detection and spatial normalisation processing steps are carried out. Can the PLSgui read in this entire timeseries or must the time volumes be somehow extracted from this file? A: For MRI module, PLSgui can read the entire timeseries volumes. For PET / Structural module, PLSgui can only read 1 volume by 1 volume. In that case, you can always use "expand_nii_scan.m" in PLSgui program to expand 1 entire timeseries volumes into 1 folder. In addition, the best way to convert AFNI BRIKs into NIfTI files is to use 3dAFNItoNIFTI. The best way to convert NIfTI files into AFNI BRIKs is to use 3dcopy. Q: I literarily counted those active voxels, why are they not listed in PLSgui cluster report? The two clusters in the cluster report are within the same cluster, why PLSgui separate them into two different clusters? The result of cluster report on one computer is totally different from the cluster report from the other one, but I have the same result file, and the same threshold. Why do I get the different results? A: In order to answer the questions, I have to describe the technique used to generate cluster report. The author of the cluster report program uses a 3D 6-connected operator to recursively flood and fill the active voxels of the cluster. The MATLAB on each computer can have different recursion limit, that's the reason you may expect different cluster report on different computers. Also because of the recursion limit, the flood and fill process may not complete, i.e. there are still active voxels in the cluster that have not been identified. That is the reason that some active voxels may not be listed in PLSgui cluster report. When cluster report program start the flood and fill from a different location, the new cluster may join one or more of the old cluster(s). In that case, all those clusters have to be combined together. However, if the new cluster can not join the old cluster(s), it will look like an individual cluster, although it could be part of the other cluster. That is the reason that PLSgui may separate active voxels in the same cluster into two or more different clusters.