landing in saccade trials, the MSD from when a target/distracter was presented in the periphery to saccadic landing was measured for each subject in a set of 70 trials during which only eye tracking data were recorded. The MSD was defined as the mean across the 70 image presentations in the periphery of the sum of the time to saccade onset after image presentation in the periphery (i.e. subject reaction time) plus the duration of the saccade before saccade landing (i.e. transit time from fixation to the peripheral target). Image presentation duration in fixation trials was 150 ms. The duration of image presentation in saccade trials was set to 150 ms plus the MSD as individually measured for each subject in order to ensure that the image presentation duration after saccadic landing was also approximately 150 ms. Image Visibility Control Procedures Both target and distracter images had Michelson contrasts relative to the black background of 0.95. The overall contrast of the images was incremented/decremented randomly between + and –5% to remove any low frequency cues to stimulus identity that subjects might be able to distinguish using their peripheral vision prior to saccade onset. The random jittering was applied both to stimuli in the saccade condition and the fixation condition. The presence or absence of the high spatial frequency rectangular grating that distinguished a target from a distracter was not detectable without making a saccade. This was validated for every subject by performing test trials prior to EEG data collection during which each subject maintained central fixation and attempted to distinguish targets from distracters without moving their eyes from central fixation when the targets/ distracters were presented in the positions 10u from center as they would be in saccade trials. It was found that all subjects were performing at chance levels in the test trials. All of the subjects also verbally confirmed that they were unable to distinguish the target and distracter stimuli in the periphery without moving their eyes from the central fixation point. EEG and Eye Movement Pre-processing EEG data were filtered with a low pass infinite impulse response (IIR) filter from the EEGlab toolbox [26] with a cutoff of 40 Hz and a high pass IIR filter with a cutoff of 0.3 Hz. Both the high and low pass filters had zero phase distortion due to forward and reverse filtering of the data. The EEG data from fixation trials were epoched using the data flanking one second (61) from the moment of image presentation (time 0). Similarly, in the case of saccadic trials, epochs were extracted and event-related averages were calculated 61 second relative to saccadic landing. Even with high pass filtering, saccades are expected to introduce step-like DC artifacts to EEG data due to the rotation of the cornea-retinal dipoles during saccades. Since the cornea retinal dipoles change positions during saccades, a different DC offset is expected before and after a sacccade [27,28]. Therefore, in addition to the highpass filter, in both fixation and saccade trials, a baseline DC level for every fixation and saccade trial was determined by averaging the DC voltage offsets associated with a 200 ms long period located from 2300 to 2100 ms before the time locking event (the time locking event was image presentation time for fixation trials and saccade landing for saccade trials) and a 100 ms long period located from 900 ms to 1000 ms after a trial’s time locking event. The mean DC voltage as determined over these pre and post windows was subtracted from each fixation and saccade trial as a baseline estimate. Note that the saccade trials had two different DC offsets in the pre and post windows defined above due to the aforementioned effects of cornea retinal dipole rotation during saccades, whereas fixation trials tended to have more The Fixation and Saccade P3 similar DC offsets in the pre and post windows. The same scheme was used for baselining for both trial types, however, in order to facilitate comparisons of the P3s observed in the fixation and saccade cases. The purpose of the aforementioned baselining procedures was to minimize the differences in DC offsets between trials prior to calculating EPs. Note that the baseline procedures did not remove step functions associated with saccades. As described in the following paragraph, saccade step artifacts were minimized in this study due to the equivalent number of leftward and rightward saccade trials. When there are approximately an equal number of leftward and rightward saccade trials with approximately equal saccade amplitudes used to calculate a saccade-locked event related average, the DC step artifacts for leftward and rightward saccades tend to cancel in the event related average due to the leftward and rightward step artifacts having approximately equal amplitudes with opposite polarities [27]. In this study, there were an equal number of leftward and rightward saccade trials with the same saccade amplitudes. Since our primary purpose was to compare the saccade and fixation P3, we therefore averaged leftward and rightward saccadic P3 responses and did not attempt to address effects pertaining to lateralization of saccades on the P3. For both trials at fixation and trials with saccades, epochs with saccades exceeding 1u following image presentation and saccade landing, respectively, were excluded from analyses. A previously described [29] microsaccade detection algorithm was used for detection of breaks from fixation. Identifying the P3 in Fixation and Saccade Trials within Subjects Standard, non-parametric cluster statistics that are commonly applied to both fMRI [30] and EEG/MEG analyses [31,32] were applied to test for significant differences between target and distracter trials in the evoked potentials (EPs) time locked to image presentation in the fixation trials and time locked to saccadic landing in the saccade trials. Cluster statistics measure the false alarm rate, account for the multiple comparisons problem, and take into account correlations across electrodes and across time. The non-parametric methods for statistical testing were applied using Fieldtrip software for EEG/MEG analysis (see http://www. ru.nl/fcdonders/fieldtrip, Donders Institute for Brain, Cognition and Behavior, Radbound University Nijmegen, the Netherlands) and are described briefly below. The first step in applying cluster statistics was to gather all of the target and distracter EEG epochs in a single condition (saccade or fixation) into a single pool of epochs. For each subject, if there were T target trials and D distracter trials, for each of 1000 iterations, T and D epochs were randomly selected from the pool of epochs of all types with disregard to whether the T and D randomly selected epochs were target or distracter trials. Target minus distracter t-statistics were determined for both the real target and distracter epochs and the 1000 iterations of randomly chosen target and distracter epochs. On each iteration, t-statistics were ‘‘clustered’’, or summed across temporally adjacent time points and spatially adjacent electrodes displaying a single sample p-value of 0.05 or less. The maximum of the sum of the clustered t- statistics was recorded on each iteration, thereby forming a distribution of t-statistic maxima observed in the randomly chosen epochs. The final p-value for any given cluster of target versus distracter difference observed in the real data could be determined by calculating the percentage of maxima of summed t-statistics in the distribution greater than the t-statistic sum associated with the cluster observed in the real data. One-tailed tests using the distributions calculated separately for saccade and fixation trials PLOS ONE | www.plosone.org 4 November 2012 | Volume 7 | Issue 11 | e48761