Near-infrared transillumination imaging combined with aperture photometry for the quantification of melanin in the iris pigment epithelium

Maciej Czepita

doi:10.1371/journal.pone.0230210

. 2020 Mar 6;15(3):e0230210. doi: 10.1371/journal.pone.0230210

Near-infrared transillumination imaging combined with aperture photometry for the quantification of melanin in the iris pigment epithelium

Maciej Czepita ^1,^*

Editor: Ireneusz Grulkowski²

PMCID: PMC7060071 PMID: 32143214

Abstract

Near-infrared transillumination is used in the diagnosis and the management of different eye diseases. In particular, it enables the visualization of melanin in the pigment epithelium of the iris. This technique is valuable in such conditions as pigment dispersion syndrome and Adie’s tonic pupil. Thus, objective quantification of the amount of melanin shedded from the iris pigment epithelium may help in the management of these conditions. By combining aperture photometry with near-infrared iris transillumination this can be achieved. A total of 4 patients (7 eyes) were examined. Three patients were diagnosed with pigment dispersion syndrome in both eyes. One patient had Adie’s tonic pupil in one eye. Near-infrared iris transillumination was performed by using a prototype apparatus. Aperture photometry measurements were carried out through specially developed software. The signal-to-noise ratio of the prototype apparatus was 52 dB (399:1). Each pixel within the near-infrared transillumination image corresponded with an area size of the iris of 85 μm x 83 μm. Measurements were taken from several points of the iris in all patients. The average aperture photometry value of transillumination defects was 1321.53 (ADU) ± 501.08 SD, while the average aperture photometry value of the papillary ruff was 90.83 (ADU) ± 53.4. On average transillumination defects transmit 14.55 times more near-infrared light than the papillary ruff. A prototype apparatus for the capture of near-infrared iris transillumination images and custom software enabling aperture photometry measurements of the obtained images has been developed for the purpose of this study. This study demonstrates a potential application of this technique in the diagnosis and management of patients with such conditions as pigment dispersion syndrome and Adie’s tonic pupil.

Introduction

Transillumination is a medical diagnostic technique of tissue illumination by transmission of light through the examined tissue [1–6]. In ophthalmology transillumination is used in the diagnosis and management of several conditions. Most notably in the localization of intraocular tumors [7], ciliary body cysts [8] and the diagnosis of pigment dispersion syndrome (PDS) [9], pigmentary glaucoma and Adie’s tonic pupil [10].

Aperture photometry is a basic image analysis technique of measuring the flux from a light source within a predefined region of interest referred to as an aperture [11]. A representation of the object’s flux is calculated by summing up all the pixel values within the aperture after subtracting an estimate of the background flux from pixel values of a background annulus centered on the aperture.

The aim of this study is to describe near-infrared iris transillumination imaging combined with aperture photometry in the quantification of melanin within the iris pigment epithelium. Due to a lack of commercially available equipment for this purpose, a prototype near infrared iris transillumination system and special aperture photometry software were developed. Special attention has been placed on measuring the amount of melanin present within the iris pigment epithelium of patients with pigment dispersion syndrome and Adie’s tonic pupil. In these conditions significant loses of melanin within the iris pigment epithelium occurs. In pigment dispersion syndrome it is currently believed that loss of melanin from the iris pigment epithelium occurs due to midperipheral posterior concavity of the iris [12]. During physiological pupillary movement, rubbing of the iris pigment epithelium against the anterior zonular bundles occurs in these patients. This leads to shedding of pigment, which gets transported with the movements of aqueous humor. The shedded pigment may deposit on the posterior capsule of the lens or on the corneal endothelium, anterior surface of the iris and also on the trabecular meshwork of the iridocorneal angle. The obstruction to the outflow of aqueous humor caused by shedded pigment on the trabecular meshwork of the iridocorneal angle leads to an increase in intraocular pressure, which in turn leads to the development of pigmentary glaucoma. Loss of the iris pigment appears clinically as midperipheral, radial, slitlike pattern of transillumination defects. These defects can be occasionally seen by retroillumination in a slit lamp or by using a transilluminator. However, near-infrared iris transillumination provides the most sensitive method of detection [13]. Quantifying the amount of melanin shedded from the iris in patients with pigment dispersion syndrome and pigmentary glaucoma has been previously done by measuring the sizes of transillumination defects from near-infrared photographic images and also by measuring the amount of melanin granules present in the aqueous humor by means of the cell count mode of the laser flare-cell meter [14,15,16]. However, no observations using aperture photometry of near-infrared iris transillumination images were carried out previously.

In the case of Adie’s tonic pupil, melanin in the iris pigment epithelium layer is believed to be shedded as a result of segmental iris atrophy [10]. These areas can be only clinically detected through iris transillumination. They are located near the pupillary margin and usually have a circular shape. Near-infrared iris transillumination provides the most sensitive method of detection. Quantifying the amount of melanin shedded has only been done by measuring the sizes of near-infrared transillumination defects [17]. No observations using aperture photometry of near-infrared iris transillumination images have been done previously.

Near-infrared iris transillumination was first used in 1977 by Saari et al. [18] in patients with Fuchs heterochromatic iridocyclitis. Following this study, Alward et al. [9] described the use of this technique in 1990 in detecting posterior iris melanin loss in patients with pigment dispersion syndrome and pigmentary glaucoma. Further studies on the potential use of this method in other conditions were later carried out [19, 20]. Modification of the equipment have also been described. Due to the properties of melanin within the iris pigment epithelium, light in the near-infrared is absorbed less than in the visible making iris transillumination defects more prominently visible. Therefore, measuring the flux of the near-infrared light within iris transillumination defects through aperture photometry enables the quantification of the amount of melanin present in the iris pigment epithelium. The areas with less melanin have a higher flux value and vice versa.

Methods

Study participants

A total of 4 patients (7 eyes) were examined. Three of the patients had pigment dispersion syndrome, while one patient suffered from Adie’s tonic pupil in one eye. Informed consent was obtained from each participant. The study followed the tenets of the Declaration of Helsinki and was approved by the Bioethics Committee of the Pomeranian Medical University. The individual pictured in Fig 1 has provided written informed consent (as outlined in PLOS consent form) to publish their image in this article.

Fig 1 — A–cooling fan. B–camera with lens. C–near-infrared transilluminator. D–near-infrared iris transillumination examination conducted by the author.

Description of prototype apparatus

The prototype near-infrared iris transillumination apparatus consists of a monochrome digital video camera sensitive to near-infrared light (WAT-910HX/RC, Watec Co., Ltd., Tsuruoka, Japan) with a macro lens (80CS20-50, 5mm, f2.2, LENEX). The camera with the lens is attached to a standard 1/4” camera tripod and placed in front of the patient on an ophthalmic table with a head and chin rest apparatus. The examination is conducted in a dark room. As a source of infrared illumination a modified MAGLITE Solitaire LED flashlight (Mag Instrument Inc., Ontario, CA, U.S.A.) is used (Fig 1). The modification involved removing the original light-emitting diode (LED) from the flashlight and replacing it with a 940 nm infrared LED diode. In order to lower the amount of thermal emission from the camera and thereby reduce the dark current of the camera a cooling fan is used (Geoptik, San Giovanni Lupatoto, Italy). The camera is placed inside the cooling fan for 10 minutes. The cooling fan is switched on. Afterwards the camera is removed from the cooling fan and connected to the tripod and objective. The patient is seated. The tip of the flashlight is gently pressed against the skin of the lower eyelid and the focus of the camera adjusted manually. A video recording of the procedure is made and stored on a laptop computer connected to the camera through a universal serial bus (USB) cable. The video is recorded for every examination with the corresponding Patient-ID-Number with SharpCap software version 3.0 (AstroSharp Ltd., Harwell, U.K.). Video is edited through Free Video to JPG Converter (Digital Wave, Ltd., London, U.K.). An exemplary near-infrared iris transillumination image can be seen in Fig 2. Aperture photometry of the images is performed using the custom made Aperture Photometry Tool for Infrared Iris Transillumination Imaging software -APTITI (2^nd Department of Ophthalmology, Pomeranian Medical University, Szczecin, Poland). The results of aperture photometry measurements are given in analog to units (ADU).

Fig 2 — (A). The pupillary ruff can be seen as a narrow dark circular band at the pupillary margin (red arrow). For comparison a visible light image of the iris of the same eye can be seen (B).

Retinal irradiance calculation

To assess the potential hazard to the retina by the infrared transilluminator of the prototype apparatus the retinal irradiance was calculated.

Retinal irradiance was calculated according to the following equation [21]:

E r = \frac{π L s τ {d e}^{2}}{{4 f}^{2}}

Er is the retinal irradiance (mW/cm²), Ls is the source irradiance (mW/cm² sr), f is the effective focal length of the eye (in centimeters), de is the pupil diameter (in centimeters) and τ is the transmittance of the ocular media. The source irradiance (Ls) of the infrared LED used was 60 mw/cm² sr. The transmittance τ of the ocular media is estimated to be 0.9 in normal patients[22]. A pupil diameter (de) of 0,7 cm was used as the examination was carried out under scotopic conditions in a dark room. The pupil dilates under these conditions to this size. The effective focal length of the eye f = 1.7 cm was used according to the model eye of Gullstrand [23]. Therefore Er in this study was:

E r = 0.27 L s τ {d e}^{2} = 7.14 m W / {c m}^{2}

According to the International Commission of Non-Ionizing Radiation Protection (ICNIRP) the retinal exposure limit to near-infrared light (IR-A radiation) is 6 W/(cm² sr) and the irradiance limit on the retina is 700 mW/cm² [24]. The calculated Er of 7.14 mW/cm² is well below the threshold value of 700 mW/cm². Therefore, no damage to the retina with the proper use of the prototype apparatus should be expected.

Compensating for defects within the CCD sensor of the camera

It is generally difficult to obtain a charge-coupled device (CCD) sensor free of image defects. Evaluating what kind of defects were present in our camera was done by cooling the camera for 10 minutes and afterwards obtaining a 10 second dark frame and a 10 second flat field image. Several hot pixels were noticed in the dark frame. These are pixels with higher than normal dark current. By superimposing the dark frame from the images obtained from our patients the exact positions of the hot pixels could be seen (Fig 3). Aperture photometry measurements using the Aperture Photometry Tool for Infrared Iris Transillumination Imaging were not performed at these locations in order not to obtain a false reading. The flat field image was recorded to spot any variabilities in sensitivities across the sensor. Variations in brightness were noticed at the edges of the flat field image due to vignetting. Dividing the near-infrared iris transillumination image pixel by pixel, by the obtained flat field image effectively removed these variations. However, through this process the connection of the pixel intensities to the collected photons became lost. Therefore, flat fielding was not applied to the obtained images. As a means to minimize the dark current of the CCD sensor of the camera we employed a thermo-electric cooler. The camera operating at room temperature was cooled by 9 degrees C.

Fig 3 — Many hot pixels can be seen (typical hot pixel marked by red arrow). No hot pixels were detected within the transillumination defects nor the pupillary ruff.

Precision of measurement

The signal-to-noise ratio of the Watec WAT 910HX/RC is 52 dB—399:1. This means that the signal is 399 times greater than the noise level or in other words the noise constitutes for 0.25% of the measurement. Therefore, the precision of the aperture photometry measurements is very high. If for example a baseline aperture photometry score is 1000 ADU, the amount of noise is equal to 2,5 ADU.

On average near-infrared iris transillumination images were captured from a distance of 4.7 cm. Given that the camera sensor is ½” and the size of the objective used is 5 mm, the horizontal width field of view of the captured images was 60 mm while the vertical height field of view of the captured images was 40 mm. This equals to a pixel size of the captured image to be 85 μm x 83 μm.

This implies that the aperture photometry measurements taken by the prototype apparatus have an accuracy of 399:1 for a minimal area size of the iris of 85 μm x 83 μm.

Results

Image processing

Once image acquisition is completed the video is exported to the Free Video to JPG Converter. The converter extracts all the frames of the video and saves them as JPEG images 720x480 pixels in size. This enables the examiner to pick out the best images for further evaluation with the Aperture Photometry Tool for Infrared Iris Transillumination Imaging-APTITI (Fig 4).

Aperture photometry measurement with APTITI

The Aperture Photometry Tool for Infrared Iris Transillumination Imaging is a custom developed plugin written by the author of this study in JAVA for ImageJ (U.S. National Institutes of Health, Bethesda, Maryland, USA) image processing software. It is available for download along with an instruction manual at https://figshare.com/projects/Aperture_Photometry_Tool_For_Infrared_Iris_Transillumination/74913. Aperture photometry measurements of the near-infrared iris transillumination images are taken by clicking on the region of interest. A set of two apertures appear on the image–an inner and outer annulus. The size of these apertures can be adjusted. With both apertures of different sizes it is possible to get a net integrated count., which is the sum of all analog to digital units (ADU) after subtracting the background values from each pixel. In order to obtain the total counts without background subtraction, the radii of the inner and outer annulus need to be set to the same value. The value of the background will be zero. The obtained result will be the total of all the analog to digital units within the aperture. This second method was used in our measurements (S1 Fig).

In this study the iris is divided into four quadrants (Fig 5). The near-infrared iris transillumination defects are identified as well as the pupillary ruff. The pupillary ruff is a small portion of the iris pigment epithelium located at the edge of the pupil of the eye. It is not affected by the disease process in pigment dispersion syndrome and Adie’s tonic pupil. It serves as a reference when performing aperture photometry. The measurements from the near-infrared iris transillumination defects are compared to the measurements from the pupillary ruff. This enables the quantification of the melanin present within the defects. An average value of the papillary ruff is calculated from measurements taken in different locations. This value is compared with the aperture photometry values of the transillumination defects. In Fig 5 (subject 1) the average aperture photometry value of the papillary ruff in the left eye is 61 ADU (Table 1). The average aperture photometry value from the upper right quadrant (A) is 2145 ADU. This implies that the transillumination defects in this quadrant transmit on average 35 times more infrared light than the pupillary ruff does. This signifies a significant loss of melanin within the iris pigment epithelium. The right eye in this patient remains not that severely affected. The average aperture photometry value from transillumination defects of all quadrants of the iris is 1226 ADU, which is only around 7,5 times higher than the papillary ruff. This difference between both eyes is most probably caused by the fact that this patient underwent a pars plana vitrectomy with silicone oil injection for a retinal detachment of the left eye. The examination was carried out 1 month after the operation. In the remaining patients such significant differences were not observed. These patients had never undergone any intraocular surgery. Overall, the average aperture photometry value of transillumination defects was 1321.53 (ADU) ± 501.08 SD, while the average aperture photometry value of the papillary ruff was 90.83 (ADU) ± 53.4. Transillumination defects transmit on average 14.55 times more near-infrared light then the papillary ruff. This information enables more precise identification of quadrants potentially at risk of greater loss of melanin form the iris pigment epithelium.

Table 1. Characteristics and results of patients examined in this study.

Subject Disease	Right eye					Left eye
Sex and age	Upper right quadrant average value in ADU	Lower right quadrant average value in ADU	Lower left quadrant average value in ADU	Upper left quadrant average value in ADU	Pupillary ruff average value in ADU	Upper right quadrant average value in ADU	Lower right quadrant average value in ADU	Upper left quadrant average value in ADU	Lower left quadrant average value in ADU	Pupillary ruff average value in ADU
Subject 1 PDS, M, 32 y.o.	1117	1042	1298	1448	164	2145	2192	2065	2107	61
Subject 2 PDS, M, 24 y.o.	1534	no defects	1247	no defects	136	1226	1427	1421	997	103
Subject 3 PDS, M, 40 y.o.	no data	no data	no data	no data	no data	no defects	885	no defects	629	21
Subject 4 Adie’s, F, 33 y.o.	no defects	no defects	no defects	no defects	no data	760	664	905	no defects	60

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PDS–pigment dispersion syndrome, ADU–analog to digital units, M–male, F—female

Discussion

The design and operation of a prototype device for near-infrared transillumination imaging of the eye has been presented along with a technique for the quantification of the amount of melanin within the iris pigment epithelium. This new technique based on aperture photometry enables more precise measurement of the amount of melanin within the iris pigment epithelium as compared to techniques used in previous studies. Instead of simply measuring the size of transillumination defects in near-infrared images of the iris as done by Haynes et al. [16] it is now possible measure the amount of melanin within each defect by using aperture photometry. This is an advantage as surely iris transillumination defects must differ from each other by the amount of melanin within.

In this study it was observed that a very low intensity infrared light-emitting diode (LED) of only 60mW/cm² sr combined with the WATEC WAT-910HX/RC camera was sufficient to obtain good quality near-infrared transillumination images of the eye. The quantum efficiency of the camera sensor at the wavelength used (940 nm) was equal to 15%. This is almost twice the quantum efficiency level at this wavelength for conventional charge-couple device (CCD) sensors. The frame rate of the during video acquisition was set to 30 fps. This enables to capture on average around 100 frames in between each blinking of the eye. As a result of the high camera sensitivity and low intensity infrared light source used the retinal irradiance was also found to be insignificant with levels reaching only 1% of the maximal allowed norm according to the International Commission of Non-Ionizing Radiation Protection (ICNIRP). However, certain disadvantages of the camera also became apparent in this study. The main problem were defective pixels. Some hot pixels were discovered in the dark frame images. In order not to compromise the aperture photometry measurements these have to be avoided during video acquisition. This required lengthy preparation of the camera and patient positioning. A lesser problem was the signal-to-noise ratio of the camera. At 52 dB the signal-to-noise ratio (SNR) is somewhat high. However, given the high sensitivity of the camera (0.000005 lux) this was acceptable.

Conclusions

A prototype apparatus for the capture of near-infrared transillumination images of the eye has been developed along with special JAVA based software for aperture photometry of the acquired images. This is the first time aperture photometry has been used in the quantification of melanin within the iris pigment epithelium. The design, operation and potential application of this new equipment have been described. The described method offering objective and reproducible quantification of melanin will allow for more accurate evaluation of the natural course and the effects of various treatment options in patients with different ocular conditions.

Supporting information

S1 Fig. Aperture photometry measurement of near-infrared iris transillumination defects using the Aperture Photometry Tool for Infrared Iris Transillumination Imaging.

(TIF)

Click here for additional data file.^{(3.4MB, tif)}

Data Availability

The software used in this study can be freely downloaded by using this link: https://figshare.com/projects/Aperture_Photometry_Tool_For_Infrared_Iris_Transillumination/74913.

Funding Statement

The author received no specific funding for this work.

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PLoS One. doi: 10.1371/journal.pone.0230210.r001

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Ireneusz Grulkowski

8 Jan 2020

PONE-D-19-27146

Near-infrared transillumination imaging combined with aperture photometry for the quantification of melanin in the iris pigment epithelium

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Reviewer #1: Dear Maciej Czepita,

Thank you very much for choosing PLOS ONE journal to submit your research about „Near-infrared transillumination imaging combined with aperture photometry for the quantification of melanin in the iris pigment epithelium „. It is a useful paper since a clinical point of view. Nevertheless, I would like to ask you questions and give you some comments in order to make clear your research:

-In the introduction, there is short description about role of the melanin in Adie’s tonic pupil (lines 85-88). The appropriate reference about it is missing.

-„A total of 4 patiens (7 eyes) were examined. A total of 4 patients (7 eyes) were examined. Three of the patients had pigment dispersion syndrome, while one patient suffered from Adies’ tonic pupil in one eye. Informed consent was obtained from each participant „ (lines 108-109). More patients should be examined. Information about age and sex is missing. The healthy patients also should be examined (as a control).

- „Description of the prototype apparatus” – there should be scheme how the whole system with sample looks like during measurement – not only single parts of the set up (Fig.1).

- How much time takes examination of the one eye? If there is some kind of adaptation to dark conditions before measurement?

- Line 134: „An exemplary near-infrared iris transillumination image can be seen in Fig 2.” Should be indicate A or B.

-Fig.2. – The scale is missing. Better is to superimpose both images.

-„The transmittance τ of the ocular media is estimated to be 0.9 in normal patients” (lines 153-154) – how did you estimate the transmittance?

- „A pupil diameter (de) of 0,7 cm was used as the examination was carried out under scotopic conditions in a dark room.”(lines 154-155) – why exactly 0,7 cm?

-„ The effective focal length of the eye f = 1.7 cm was used according to the model eye of Gulstrand.” (lines 156-157) – the reference is missing

- „By superimposing the dark frame from the images obtained from our patients the exact positions of the hot pixels could be seen (Fig 3).” (lines 173-175) There should be arrows to indicate examples of hot pixels.

- „precision of measurements”, line 195, please explain in the text, what ADU means.

- Table – 1, please explain in the text, what PDS means.

Reviewer #2: The following old and recent papers on transillumination for different applications should be considered in the introduction:

DOI: 10.1109/10.817628

DOI: 10.1364/OL.15.001179

DOI: 10.1109/TBME.2003.812188

DOI: 10.3390/s19040851

DOI: 10.1016/j.biosystemseng.2014.06.014

DOI: 10.1016/j.compag.2019.02.014

Usually transillumination refers to a setup in which light source and detector are one in front of the other. In this case light source and detector are at 90°. Would you be able to comment on the differences? Are you actually detecting side scattered photons? Or is there another way to describe the phenomenon?

Why CCD and not a CMOS? There are CMOS sensors with enhanced responsitivy in the NIR.

Did you need dark room to ensure that the pupil is dilated or to avoid camera saturation with ambient light? In this second case, a narrow optical filter in front of the camera should be sufficient to select only the 940 nm photons

You need to define the acronyom ADU (Analog to Digital Unit) and explain the meaning of it in your experiments.

In many places the authors claim that their measurements enable the quantification of the

melanin. But I do not see any results reporting the exact amount of melanin. Only ADU values are reported but they are not indicating the amount of melanin. So the authors should solve this issue.

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PLoS One. 2020 Mar 6;15(3):e0230210. doi: 10.1371/journal.pone.0230210.r002

Author response to Decision Letter 0

29 Jan 2020

Response to remarks of Reviewer #1

1. I have now included an appropriate citation regarding the role of melanin in Adie’s tonic pupil (line 85-88).

2. I have now included information about the patients age and sex in Table 1. The overall incidence of pigment dispersion syndrome is thought to be around 4.8 per 100,000 population/year1. The overall incidence of Adie’s tonic pupil is approximately 4.7 per 100,000 population/year2. Both conditions are rare. The 4 patients participating in our study were recruited over a 2 year period. One patient newly diagnosed with pigment dispersion syndrome did not wish to participate in the study. A future study on a larger group of patients will be undertaken. The main aim of this paper is to demonstrate the feasibility of this technique in these conditions. A control group was not examined in this study as the transillumination defects are not present in healthy individuals. Therefore, there wouldn’t be anything to compare among the two groups.

3. I have now included an additional photograph in Figure 1 displaying the setup of the experimental apparatus during examination.

4. The examination takes around 1 minute per patient. Dark room adaptation before the examination was 1 minute.

5. I have now added indicated the A and B subpart in Figure 2.

6. I have added a scale in both subparts of Figure 2. It is not possible to superimpose both images of the iris in the visible and near-infrared because the pupil is larger in the near-infrared than in the visible. This is due to pupil dilatation in dark conditions and pupil constriction in light.

7. I have included now a citation on regarding the transmittance of the ocular media (line 153-154)

8. In the equation (line 149) we assumed a pupil diameter of 7 mm as this is the mean value for the examined age group.

9. The citation for the effective focal length of the schematic eye of Gullstrand has now been included (lines 156-157)

10. I have now added a red arrow in Figure 3 pointing to a hot pixel. I have added a explanation on ADU in the description of the prototype apparatus section of the manuscript.

11. I have now added an explanation of PDS in the manuscript.

References:

1.Ritch R, Steinberger D, Liebmann JM. Prevalence of pigment dispersion syndrome in a population undergoing glaucoma screening. Am J Ophthalmol. 1993 Jun 15;115(6):707-710.

2. Sarao MS, Sandeep S. Adie Syndrome https://www.ncbi.nlm.nih.gov/books/NBK531471/

Response to remarks of Reviewer #2

1. I have added in the introduction citations about the various medical uses of NIR transillumination.

2. The near-infrared transilluminator is held at an angle of around 60 degrees to the camera during the examination in order for the light to penetrate through the sclera and vitreous chamber and then reflect of the retina. The reflected light then causes backlighting of the iris. Therefore, the iris pigment epithelium can be imaged through this technique.

3. The WATEC WAT 910-HX/RC CCD camera was chosen because of it’s enhanced sensitivity in the NIR as well as the price which was within our budget for the study.

4. The examinations were carried out in a dark room in order avoid camera saturation with ambient light

5. The acronym ADU is now explained at first in the description of the prototype apparatus section of the manuscript.

6. In the study I compared the aperture photometry readings of the transillumination defects with the aperture photometry readings of the papillary ruff. The papillary ruff is the portion of iris pigment epithelium at the margin of the pupil. This part of the iris pigment epithelium is not affected by the disease process in pigment dispersion syndrome and usually also in Adie’s tonic pupil. By comparing the both results the amount of melanin shedded it can be estimated.

PLoS One. doi: 10.1371/journal.pone.0230210.r003

Decision Letter 1

Ireneusz Grulkowski

25 Feb 2020

Near-infrared transillumination imaging combined with aperture photometry for the quantification of melanin in the iris pigment epithelium

PONE-D-19-27146R1

Dear Dr. Czepita,

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PLoS One. doi: 10.1371/journal.pone.0230210.r004

Acceptance letter

Ireneusz Grulkowski

27 Feb 2020

PONE-D-19-27146R1

Near-infrared transillumination imaging combined with aperture photometry for the quantification of melanin in the iris pigment epithelium

Dear Dr. Czepita:

I am pleased to inform you that your manuscript has been deemed suitable for publication in PLOS ONE. Congratulations! Your manuscript is now with our production department.

If your institution or institutions have a press office, please notify them about your upcoming paper at this point, to enable them to help maximize its impact. If they will be preparing press materials for this manuscript, please inform our press team within the next 48 hours. Your manuscript will remain under strict press embargo until 2 pm Eastern Time on the date of publication. For more information please contact onepress@plos.org.

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Associated Data

This section collects any data citations, data availability statements, or supplementary materials included in this article.

Supplementary Materials

S1 Fig. Aperture photometry measurement of near-infrared iris transillumination defects using the Aperture Photometry Tool for Infrared Iris Transillumination Imaging.

(TIF)

Click here for additional data file.^{(3.4MB, tif)}

Data Availability Statement

The software used in this study can be freely downloaded by using this link: https://figshare.com/projects/Aperture_Photometry_Tool_For_Infrared_Iris_Transillumination/74913.

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Near-infrared transillumination imaging combined with aperture photometry for the quantification of melanin in the iris pigment epithelium

Maciej Czepita

Roles

Abstract

Introduction

Methods

Study participants

Fig 1. Main components of prototype apparatus.

Description of prototype apparatus

Fig 2. Near-infrared iris transillumination image of the left eye of a healthy volunteer.

Retinal irradiance calculation

Compensating for defects within the CCD sensor of the camera

Fig 3. Result of superimposing a dark frame image with a near-infrared iris transillumination image.

Precision of measurement

Results

Image processing

Fig 4. Flowchart of image analysis algorithm.

Aperture photometry measurement with APTITI

Fig 5. Near infrared iris transillumination image of a patient with pigment dispersion syndrome (subject 1).

Table 1. Characteristics and results of patients examined in this study.

Discussion

Conclusions

Supporting information

Data Availability

Funding Statement

References

Decision Letter 0

Ireneusz Grulkowski

Roles

Author response to Decision Letter 0

Decision Letter 1

Ireneusz Grulkowski

Roles

Acceptance letter

Ireneusz Grulkowski

Roles

Associated Data

Supplementary Materials

Data Availability Statement

ACTIONS

PERMALINK

RESOURCES

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