positions, the number of pixels in the transverse direction is equal to the number of axial scans. A typical OCT retinal image is 6 mm wide in transverse direction and has 512 transverse pixels, so the pixels are 6 mm/512 ¼ 11.7 mm wide. In contrast, the transverse resolution for a typical retinal image is determined by the focused spots size and is typically 20 mm. The pixel density in the axial direction is determined by the speed at which the computer can record the electronic signal from the axial scan of backscattering or back reflection versus depth. The speed can be very rapid. The typical OCT retinal image is 2 mm deep in axial direction and acquires 1024 pixels, therefore the pixel size is 2/1024 ¼ 1.9 mm in axial depth. Finally, in order to utilize the full instrumental resolution, the size of the pixel must be smaller than the instrument resolution.

The speed of image acquisition is another important factor. Rapid acquisition not only minimizes image distortion and artifact, but also improves patient comfort. The speed is directly related to the sensitivity of the measurement because imaging more rapidly reduces signal-to-noise performance. This performance could be improved using higher incident optical power, but has to be limited due to safety standards. In nonophthalmic imaging applications, higher incident optical powers can be used to dramatically increase the acquisition speed. Image acquisition time also increases in proportion to the number of transverse pixels in an image. If a higher transverse pixel resolution is desired, then more axial scans are required and the acquisition time increases proportionally. Conversely, if only low-transverse resolution imaging is necessary, then the number of transverse pixels may be reduced and the image acquisition time will be proportionally decreased. The trade-off between pixel density and acquisition speed is important for imaging protocols. Commercially available OCT has an axial scan repletion rate of 400 axial scans per second. The high transverse pixel density image has 512 transverse pixels and is 6 mm wide, with an acquisition speed of 512/400 ¼ 1.28 s. The same OCT having a low-transverse pixel density with 128 transverse pixels has a speed of 128/400 ¼ 0.32 s.

In conclusion, a larger number of transverse pixels are desirable to improve the visualization of

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retinal features; however, this high-density image requires a longer time and therefore artifacts from eye motion can increase. A more precise registration is possible with faster image acquisition, but the image will appear grainier because of a reduced transverse pixel density.

How OCT is performed

Pupillary dilatation is recommended before glaucoma assessment using the Stratus OCT. Studies found that the dilated scans were more reproducible and of higher quality than the undilated scans (Smith et al., 2007).

The operator must choose among different scanning protocols to perform RNFL or optic nerve head (ONH) OCT. Commercially available OCT machines have various scan protocols. They are composed of two basic types of scans: line and circle. Each protocol is composed of line or circle scans with different parameters (number, angle, length, and diameter). Once the protocol is selected, the patient is positioned behind the instrument and the examiner is provided with a video camera to view the scanning probe beam on the fundus. At the same time, a computer monitor shows the OCT image acquired in real time. It is possible to center the circular scan on the optic nerve head while the subject fixates with the eye being studied (internal fixation technique). The centering technique depends on the examiner’s ability to perform fine positioning of the circular scan. After the image is acquired, various analysis protocols are available to represent RNFL or ONH OCT images.

Evaluation of RNFL thickness

Evaluation of the RNFL thickness is essential for diagnosing and managing glaucoma. The RNFL comprises ganglion cell axons, neuroglia, and astrocytes (Radius and Anderson, 1979; Varma et al., 1996). The ophthalmoscopic appearance of the RNFL was first described by Vogt in 1913 (Vogt, 1913). In the early seventies, numerous studies (Hoyt and Newmann, 1972; Hoyt et al., 1973) emphasized the importance of examining the

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RNFL in studying glaucoma. These studies suggested that, in some cases of glaucoma, RNFL atrophy is primarily related to a degeneration of the ganglion cell axons, followed by a thinning of the NFL. Thus, when attempting to detect early glaucomatous optic nerve damage, it would be useful to obtain objective quantitative measurements of the thickness of the RNFL in addition to qualitative observation of the defects in the RNFL.

RNFL can be assessed subjectively with a slit lamp and a handheld lens, but this technique requires experience and offers only qualitative data that are difficult to compare over time. Several instruments such as scanning laser ophthalmoscopy (SLO), scanning laser polarimetry (SLP), and OCT allow objective and quantitative evaluation of RNFL.

Nerve fiber layer thickness is assumed to be correlated with the extent of the red, highly reflective layer at the vitreoretinal interface (Fig. 1). Boundaries are located by searching for the first points on each scan where the reflectivity exceeds a certain threshold. The posterior margin of the nerve fiber layer is located by starting within the photoreceptor layer and searching upward in the image (Schuman et al., 1996).

Using the most commercially available OCT (Stratus OCT 3, Carl Zeiss Meditec Inc., Dublin, California), two protocols are available for managing glaucoma. The RNFL thickness (3.4) protocol is designed to acquire three circle scans of diameter of 3.4 mm around the optic disc. The fast RNFL

protocol compresses the three RNFL thickness (3.4) circle scans into one scan to simplify the process and shorten the acquisition time.

OCT acquires images in cylindrical sections surrounding the optic disc. Computer image processing allows for an estimation of RNFL thickness from circumpapillary scans: the RNFL thickness is defined as the number of pixels between the anterior and the posterior edges on the RNFL identified by an edge detection algorithm.

The protocol used most to study RNFL thickness is the RNFL thickness average analysis protocol (Fig. 2). The NFL thicknesses are reported as averages over either each quadrant or clock hour and as a graph of NFL thickness along the scan line (Schuman et al., 1995, 1996).

The RNFL thickness average analysis protocol is extremely useful in comparing the measurements of the NFL between the eyes. The RNFL thickness map analysis protocol is used to obtain a map of NFL thickness of the peripapillary area. To assess changes in NFL thickness between examinations, ‘‘RNFL thickness change’’ and ‘‘RNFL thickness serial’’ analysis protocols are useful.

Several studies evaluated the optimal circle diameter for OCT NFL scans based on reproducibility of measurements and disc size. Fixation technique (internal or external) and its effect on the reproducibility of OCT measurements have been evaluated. The standard circle diameter selected is 3.4 mm, which is ideal for several reasons. It is large enough to avoid overlap with the optic nerve head