Qinmei Wang, MD Affiliated Eye Hospital of Wenzhou Medical College 270 West Xueyuan Road Wenzhou Zhejiang 325027 China Tel: + 86 577 88068862 Fax: + 86 577 88832083 Email: email@example.com or firstname.lastname@example.org
Purpose: To compare the repeatability and reproducibility of central corneal thickness (CCT) measurements by high-resolution (HR) rotating Scheimpflug imaging and Fourier-domain optical coherence tomography (FD-OCT). CCT measurements were compared to those determined by ultrasound pachymetry (UP).
Methods: In 35 healthy eyes, intra-observer repeatability for HR Scheimpflug (Pentacam) and FD-OCT (RTVue) systems was determined in consecutive images taken by an observer in the shortest time possible. Imaging was repeated again by a second observer to evaluate inter-observer reproducibility. The CCT measurements were compared among Scheimpflug, FD-OCT and UP images.
Results: Mean coefficients of repeatability were 0.48% for Scheimpflug and 0.26% for FD-OCT. For Scheimpflug, the coefficient of inter-operator reproducibility was 0.87%. For FD-OCT, the coefficient of inter-operator reproducibility was 0.45%. The CCT measurements by Scheimpflug, OCT and UP images were (mean ± standard deviation) 521.7 ± 27.6 μm, 510.8 ± 28.6 μm and 516.5 ± 27.6 μm, respectively. The differences between instruments were statistically significant. The 95% limits of agreement in CCT were −0.7 to 22.5 μm for Pentacam-OCT, −13.4 to 24.0 μm for Pentacam-UP and −26.7 to 15.4 μm for OCT-UP. There was a high degree of correlation between CCT measured by all 3 methods.
Conclusion: Noncontact measurements of CCT with HR Scheimpflug and FD-OCT systems yielded excellent repeatability and reproducibility and can be used interchangeably. Although both devices were comparable with UP; in clinical practice, the measurements acquired by optical modalities are not directly interchangeable with UP measurements.
Accurate and reliable determination of corneal thickness has become increasingly necessary for the diagnosis of corneal diseases and therapeutic applications such as monitoring corneal oedema and endothelial function (Gordon et al. 2002; Ou et al. 2002; Martin et al. 2007, 2008). Currently, ultrasound pachymetry (UP) is the most frequently used clinical technique and the gold standard to evaluate central corneal thickness (CCT) (Marsich & Bullimore 2000; Miglior et al. 2004). However, UP has several possible sources of error. Its accuracy depends on the placement of the probe on the cornea, and the perpendicularity of the probe with respect to the cornea is often difficult to ascertain. If the probe is placed slightly off centre at an oblique incidence, the thickness may be overestimated. Inaccurate results can also occur after instillation of topical anaesthesia that produces epithelial oedema (Nam et al. 2006). In contrast, excessive pressure of the probe on the corneal surface and displacement of the tear film can lead to underestimation of the true thickness. Moreover, the accuracy may be influenced by variability in the speed of ultrasound in tissues of different hydration (Gonzalez- Meijome et al. 2003; Nemeth et al. 2006).
These limitations have led to a number of new and more sophisticated techniques that provide rapid, convenient, noncontact and objective measurements of the CCT. The rotating Scheimpflug imaging has become popular for clinical evaluation of the ocular anterior segment (Chen & Lam 2009). It can measure the thickness of the cornea at any point, providing elevation maps of both the anterior and posterior corneal surfaces. The latest high-resolution (HR) Scheimpflug camera has a redesigned optic design and is improved to maximally capture 138 000 data points in less than 2 seconds. The original device provides CCT measurements of normal corneas that are reliable and have high agreement with UP (Barkana et al. 2005; Lackner et al. 2005; O’Donnell & Maldonado-Codina 2005; Khoramnia et al. 2007; de Sanctis et al. 2007a,b; Shankar et al. 2008). To our knowledge, however, there are no reports on the repeatability and reproducibility of CCT by the HR rotating Scheimpflug imaging.
More recently, improvements in optical coherence tomography (OCT) technology have been introduced. The three-dimensional high-resolution OCT uses Fourier-domain (FD) detection to acquire the image much faster, which enables higher definition, less motion error and greater scanning area (Wojtkowski et al. 2005; Liu & Brezinski 2007). Typically, a commercial Fourier-domain optical coherence tomography (FD-OCT) instrument has scan rates of about 26 000 A-scans per second, 65 times faster than time domain OCT (TD-OCT) instruments. The axial resolution is 5 μm in tissue, which is a twofold improvement over TD-OCT. With a corneal adaptor module (CAM), FD-OCT obtains high-resolution cross-sectional images of the cornea and provides pachymetry. It can also provide sophisticated goniometry of the irido-corneal angle and other anterior segment structures. FD-OCT yields highly repeatable and reproducible measurements of retinal nerve fibre layer thickness (Gonzalez- Garcia et al. 2009). Because FD-OCT was designed to image the retina, it is essential to quantify the repeatability and reproducibility for corneal thickness.
The purpose of this study was to evaluate repeatability and reproducibility of CCT measurements with the HR rotating Scheimpflug imaging system and the FD-OCT system. We also compared the level of agreement with measurements obtained by UP.
Materials and Methods
In this prospective study, CCT of 35 healthy volunteers (21 men and 14 women) with a mean age of 23.9 ± 2.7 (standard deviation, SD) years (range, 18 to 29 years) was determined between November and December in 2008. The research protocol adhered to the tenets of the Declaration of Helsinki and was approved by the Office of Research Ethics, Wenzhou Medical College. Each subject gave informed consent after the nature, and intent of the study had been fully explained. The exclusion criteria were previous ocular surgery, ocular pathology, such as keratoconus or glaucoma, and contact lens wear. All measurements were performed in the undilated right eye.
We used the latest HR rotating Scheimpflug imaging system (Pentacam HR; Oculus, Wetzlar, Germany). The subject was asked to open both eyes and fixate straight ahead on a target. The room lights were switched off for all examinations to get a reflex-free image. To reduce operator-dependent variables, the automatic release mode was used. In less than 2 seconds, the rotating camera captured 25 slit images of the anterior segment. Each slit image consisted of 1 380 true elevation points that were analysed by Software 6.02r23. Only scans with an instrument-generated quality factor greater than 95% were chosen for analysis. The apex value of corneal thickness was recorded as the CCT (Shankar et al. 2008).
The high-speed FD-OCT system (RTVue-100; Optovue Inc, Fremont, CA, USA) used a near-infrared, low-coherence, super luminescent diode light source with a 50 nm bandwidth and a centre wavelength of 830 nm. It had an axial resolution of 5 μm in tissue and a transverse resolution of up to 1.5 μm. A CAM was mounted on the probe to focus the OCT beam onto the cornea. A 6-mm radial pachymetry scan pattern, centred at the corneal apex was used to map corneal thickness. It consisted of eight equally spaced meridians, and each had 1024 A-scans. To ensure that the FD-OCT scanned the central location, all subjects were asked to stare at a fixation target within the OCT system. By placing the apex at the centre of the OCT image and maximizing the apex reflection, the scanning axis became orthogonal to the corneal surface, ensuring that the observation and scanning axes were coaxial. Under these conditions, a clear reflection was obtained on a monitor, and the OCT image was recorded. The reading displayed in the centre was used for analysis. The CCT was automatically calculated by the software currently provided by the manufacturer (RTVue-100 version 3.6; Optovue Inc., Fremont, CA, USA). To reduce the speckle noise, the study images were averaged from 16 scans (Liu et al. 2008).
In the first experiment, the reliability of the rotating Scheimpflug imaging and FD-OCT was determined based on the definitions adopted by the British Standards Institution, as recommended by Bland and Altman (Bland & Altman 1986; Institution 1994a,b). The right eye CCT of each subject was measured using both methods by two independent examiners. The order of the devices with which the measurements were taken was randomized. Two scans were performed by the first examiner (HJH) and then a further scan by the second examiner (WDZ). After each acquisition, the device was moved backwards and realigned for the next scan to eliminate interdependence of the successive measurements. The time for the instrument to calculate the data between successive scans was approximately 20 seconds and 10 seconds for the rotating Scheimpflug camera and FD-OCT system, respectively. The total time for acquiring all measurements did not exceed 10 min.
In the second experiment, we compared the accuracy of CCT measurements using rotating Scheimpflug camera imaging, FD-OCT, and by UP. At the first session, after performing noncontact examinations using the rotating Scheimpflug camera system and FD-OCT system, the cornea was anaesthetized with 0.5% proparacaine hydrochloride (Alcaine; Alcon Laboratories, Fort Worth, TX, USA). The A-scan UP instrument (SP-3000; Tomey Inc., Nagoya, Japan) was precalibrated for all measurements. The ultrasound velocity was set at 1640 m/s. A handheld probe was aligned as perpendicular as possible on the central cornea. Five readings were obtained, and the highest and the lowest values were excluded. The mean of the remaining 3 was calculated and compared with the mean CCTs made by the first examiner using rotating Scheimpflug imaging and FD-OCT.
All data were analysed using the Statistical Package for Social Sciences for Windows version 13.0 (SPSS Inc, Chicago, IL, USA). Results are presented as means ± SDs. Intra-observer repeatability was calculated with the two measurements obtained by the first examiner during the first visit. The coefficient of repeatability (CoR1) and intra-class correlation coefficients (ICCs) were calculated. As recommended by Bland and Altman and proposed by the British Standard Institution (Bland & Altman 1986; Institution 1994a,b), CoR1 obtained from the repeated administration of the test under identical conditions was defined as the SD of the difference from the mean of these repeat measurements divided by the mean response. An ICC of 0.75 determined on the basis of analysis of variance for two-way mixed-effects model with an absolute agreement for consistency of individual measurements indicates good reliability. However, most clinical applications require a value of at least 0.90 (Portney & Watkins 2000).
For each of the noncontact methods, differences between the first measurements obtained by the two examiners during the same visit were used to assess inter-observer reproducibility. The inter-observer coefficient of reproducibility (CoR2) and ICC were then calculated. CoR2 was defined as the SD of the difference between measurements obtained during repetition of the test under different conditions, i.e. different observers, divided by the average of the means of each pair of readings. The normality of all data distributions was confirmed by the Kolmogorov-Smirnov test. Comparison of the mean CCT values for the three instruments was conducted by repeated-measures analysis of variance (anova). Pairwise differences were determined using the Bonferroni adjustment for multiple comparisons. The linear correlation between measurements (Pearson’s coefficient of correlation) with values >0.7 was considered indicative of good correlation between the methods. A p-value <0.05 was considered statistically significant, and 95% limits of agreement (LoA) were defined as the mean difference ± 1.96 SD of the differences. Bland–Altman plots also were used to assess the reliability of the measurements.
The mean spherical equivalent refraction in the 35 eyes was −4.01 ± 2.45 diopters.
Repeatability of central corneal thickness measurements
The CCT values measured by HR rotating Scheimpflug imaging and by FD-OCT were normally distributed (p > 0.05). Measurements of CCT were highly repeatable with both imaging systems (Table 1). For rotating Scheimpflug imaging and FD-OCT, the CoR1 was 0.48 and 0.26, respectively. The ICC for the rotating Scheimpflug imaging was 0.987 and that for FD-OCT was 0.997. Bland–Altman analysis showed that the difference between the first and second measurements was evenly dispersed around the mean, with no clear trend towards over or underestimation by either rotating Scheimpflug imaging or FD-OCT (Fig. 1). The 95% LoAs were −9.7 to +8.2 μm for rotating Scheimpflug imaging and −5.0 to +4.0 μm for the FD-OCT (Table 1). Thus, the variability between first and second measurements was larger with rotating Scheimpflug imaging than with FD-OCT, although both instruments produced reliable measurements from single examiners.
Table 1. Intra-observer repeatability of the Pentacam rotating Scheimpflug camera high-resolution system and RTVue Fourier-domain optical coherence tomography system in measuring central corneal thickness (n = 35).
Mean Difference ± SD (μm)
CoR1 (%) (95% CI)
ICC (95% CI)
SD = standard deviation, CoR1 = coefficient of repeatability, CI = confidence interval, ICC = intra-class correlation coefficient, LoA = limits of agreement according to Bland–Altman method.
−0.71 ± 4.56
−0.51 ± 2.29
Reproducibility of central corneal thickness measurements
Measurements of CCT were highly reproducible with both imaging systems (Table 2). For rotating Scheimpflug imaging and FD-OCT, the respective values of CoR2 were 0.87% and 0.45%, values of ICC ranged between 0.985 and 0.996. The inter-observer 95% LoA was −9.5 to 9.2 μm for rotating Scheimpflug imaging and −5.4 to 4.8 μm for FD-OCT (Fig. 2). Thus, the variability between examiners was larger with rotating Scheimpflug imaging than with FD-OCT, although both instruments had comparable reproducibility for CCT measurements.
Table 2. Inter-observer reproducibility of the Pentacam rotating Scheimpflug camera high-resolution system and RTVue Fourier-domain optical coherence tomography system in measuring central corneal thickness (n = 35).
Mean Difference ± SD (μm)
ICC (95% CI)
SD = standard deviation, CoR2 = coefficient of reproducibility, CI = confidence interval, ICC = intra-class correlation coefficient, LoA = limits of agreement according to Bland–Altman method.
−0.14 ± 4.78
−0.26 ± 2.60
Agreement of central corneal thickness measurements
CCTs measured with rotating Scheimpflug imaging and FD-OCT were 521.73 ± 27.62 μm and 510.83 ± 28.55 μm, respectively. CCT measured by UP was 516.46 ± 27.59 μm. There were significant variations in the CCT measurements obtained using the three methods. The CCT determined by rotating Scheimpflug imaging was significantly higher than that determined by UP (p = 0.007, Table 3). In contrast, the FD-OCT measurements were on average 5.63 μm lower than those determined by UP (p = 0.012, Table 3). The Bland–Altman plots revealed that rotating Scheimpflug imaging significantly overestimated the CCT by 10.90 μm compared with FD-OCT (p < 0.001, Table 3 and Fig. 3), as 95% of the differences in the readings between t rotating Scheimpflug imaging and FD-OCT lay between −0.7 μm and 22.5 μm. Among the three instruments, there were high linear correlations in pairwise scatter plots (p < 0.001).
Table 3. Paired t test, correlation values and limits of agreement (LoA) between 3 pairs of devices.
Mean Difference ± SD (μm)
95% LoA (μm)
UP = Ultrasound pachymetry, SD = Standard deviation, *2 tailed.
Pentacam – RTVue
10.90 ± 5.93
Pentacam – UP
5.27 ± 9.55
RTVue – UP
−5.63 ± 10.75
In using an instrument or technique for ophthalmic examination, particularly for longitudinal studies, it is important to assess its precision (Zadnik et al. 1992). If one method has poor precision, the agreement between it and other methods is bound to be poor too. It is not possible to reliably achieve accuracy in individual measurements without precision (Bland & Altman 1986). This evaluation is fundamental in assuring the predictive value of a measurement. The clinical utility of any instrument also depends strongly on the precision of its measurements. Better precision improves the ability of the clinician or researcher to detect change in a patient’s status because of the passage of time or to an intervention (Bailey et al. 1991).
In the present study, we show that repeatability and reproducibility were high with both noncontact methods, thereby allowing us to assume that these measurements may be safely carried out by different examiners and in different locations. FD-OCT had a lower (better) coefficient of repeatability and also a lower coefficient of reproducibility with narrower 95% LoA compared with the rotating Scheimpflug imaging.
Previous studies showed that the original rotating Scheimpflug imaging provides reliable CCT data in normal, post-laser in situ keratomileusis and keratoconus corneas, as well as in corneal grafts (Lackner et al. 2005; de Sanctis et al. 2007a,b; Ciolino et al. 2008). A recent study assessed the precision of CCT measurements obtained with this instrument in normal eyes. The ICCs for intra-observer and inter-observer variability were 0.99 and 0.98 for the Pentacam, the 95% LoA of repeated measurements was −12.5 to 12.5 μm, between examiners were −17.8 to 12.6 μm (de Sanctis et al. 2007a,b). (Lackner et al. 2005) found similar repeatability, and (Barkana et al. 2005) also reported good reproducibility. In contrast, (O’Donnell & Maldonado-Codina 2005) found relative larger inter-session variability with a 95% LoA of −24.1 to 21.1 μm. This may be an indication of diurnal variation in corneal thickness between different sessions. Using the HR rotating Scheimpflug system here, we report a similar high ICC values, ranging from 0.977 to 0.987. Our 95% LoAs of intra-observer and inter-observer values with the HR rotating Scheimpflug system, −9.7 to 8.2 μm and −9.5 to 9.2 μm, respectively, were narrower than with the original Scheimpflug system. The findings of our study compare favourably with previous studies (Barkana et al. 2005; Lackner et al. 2005; O’Donnell & Maldonado-Codina 2005; de Sanctis et al. 2007a,b).
The rotating Scheimpflug camera acquires images during a 180° rotation around the optical axis so that each section runs through the corneal apex. The centre of the cornea is detected automatically, and measurement alignment occurs automatically and does not rely upon the examiner. However, the patient’s correct gaze is important and can affect the precision of this method. For this study, the ‘25 images per scan’ option was chosen. The long acquisition time of about 2 seconds may be difficult for patients who must maintain a steady fixation and widely opened eye.
Another potential explanation for the differences between the precision of the original and the new HR rotating Scheimpflug imaging may be the different sampling modes of both devices. The original system captured 25 or 50 Scheimpflug slit images within 2 seconds. The images contained from 500 to 25 000 points. The new HR version is equipped with a 1.45 megapixel camera and can maximally capture 138 000 points, 5.52 times better than original version. The large number of points measured in the central area with the mapping protocol may compensate for the theoretical variability because of changes in corneal thickness with the impact of unstable fixation.
An important factor that may influence the reliability of pachymetry readings for all of the instruments is the difference in locating the measurement region. In theory, the CCT is defined as the radial distance between two concentric spheres of the cornea. However, eyes are not perfectly formed, and the anterior and posterior surfaces are not concentric, and the anterior surface is aspheric.42 Therefore, several possible ‘definitions’ of CCT exist. For example, CCT could be defined at the fixation point, visual axis, corneal pupil and thinnest region of the apex among others. In our study, the CCT refers to the apex of the cornea. Recent studies have investigated the reliability of central and peripheral corneal thickness measurements using the original rotating Scheimpflug camera system. (Khoramnia et al. 2007) found good reliability for pachymetry measurements but the reliability decreased slightly towards the peripheral areas. The mean thicknesses of the apex and the thinnest portion of the cornea were 4.33 μm and 4.49 μm, respectively. The measurements increased to 8.31 μm at 3.0 mm from the pupil. Similarly, Shankar et al. reported that CCT was comparable at pupil centre and corneal apex, but peripheral repeatability was much better when centred on the corneal apex than at pupil centre (Shankar et al. 2008). This may be explained by the measurement principle used by the Pentacam device. The number of points measured decreases from the centre to the periphery of the cornea, where the radial lines are spaced further apart. Thus, more interpolation of data is required, leading to greater variability of measurements in the peripheral corneal area. In addition, variability in pupil diameter and decentration affected peripheral pachymetry. Thus, Shankar et al. recommend that the corneal apex CCT be used for clinical purposes (Shankar et al. 2008).
To our knowledge, this is the first reported assessment of the repeatability, reproducibility and agreement of CCT measurements between the HR rotating Scheimpflug imaging and FD-OCT. We found that CCT measurements obtained by FD-OCT were highly correlated with those derived from rotating Scheimpflug imaging and from UP. The FD-OCT CCT measurement was 5.6 μm less than that determined by UP. In contrast, the CCT obtained by the HR rotating Scheimpflug imaging was 5.3 μm more than the UP value. Although the working principle of the noncontact devices is different, both instruments demonstrated similar spans of the 95% LoAs with UP as demonstrated in the Bland–Altman plots. The mean difference between both noncontact devices and UP may not be clinically significant, but the variability expressed by the 95% LoA values is broad. This suggests that clinicians should be aware of the differences between noncontact devices and UP and that they cannot be used interchangeably for clinical or research purposes.
(Ishibazawa et al. 2008) demonstrated that the FD-OCT provided CCT measurements that were highly correlated with but systematically lower than the measurements provided by UP. The mean difference was 34 μm in 30 normal subjects. Here, we found that the FD-OCT CCT measurement was slightly less than UP, with a mean difference of 5.6 μm. There are several theoretical explanations for the discrepancy between both optical methods and UP in measuring the CCT. These include decentration, oblique incidence of the probe to the cornea, possible effect of topical anaesthesia with contact pachymetry and the variability of ultrasound speed in tissues of different hydration (Gonzalez- Meijome et al. 2003; Nemeth et al. 2006; Paul et al. 2008). The accuracy of UP is operator dependent. (Reader & Salz 1987) reported a maximum difference of 49 μm in CCT measurements with different ultrasound pachymeters. For both rotating Scheimpflug imaging and FD-OCT, they calculated the distance between the anterior and posterior corneal surfaces and transformed it into corneal pachymetry. These two systems yield highly reliable measurements of corneal thickness in healthy eyes,17−20, 47 although they can be affected if the cornea is not clear or if there is other optical interference (Matsuda et al. 2008). The differences we and others report between the optical methods and UP in different research settings may be mainly attributed to sources of measurement variability inherent in UP itself.
The HR rotating Scheimpflug imaging measurement of CCT was 10.9 μm greater than that determined by FD-OCT. The 95% LoA suggested that in 95% of the cases, measurements taken by rotating Scheimpflug imaging will differ from FD-OCT measurements by less than 11.6 μm. The difference was small and comparable to the reported diurnal CCT fluctuation (Lattimore et al. 1999). It seems that for most practical purposes, measurements with both instruments can be used interchangeably.
In conclusion, the HR rotating Scheimpflug imaging and FD-OCT system yielded excellent repeatability and reproducibility for CCT measurements. While FD-OCT had a better reliability than HR rotating Scheimpflug imaging, they can be used interchangeably for measurements in normal eyes. In addition, both optical methods demonstrated good agreements with UP. But it is important to note that in clinical practice, measurement values are not directly interchangeable. Further work is required to assess the accuracy and precision of both optical methods when applied to abnormal corneas, such as keratoconus cornea, postlaser in situ keratomileusis cornea and others.
Proprietary interests: David Huang receives patent royalty, stock options, travel support and grant support from Optovue, and patent royalty from Carl Zeiss Meditec, Inc. We thank Britt Bromberg, PhD, Xenofile Editing, for providing editing services for this manuscript.