Two-photon microperimetry

Microperimetry is a subjective visual field testing method that enables the assessment of retinal function at various specific and focal locations. Two-photon microperimetry is an extension of this technique. In contrast to traditional microperimetry, which uses a visible stimulus, two-photon microperimetry utilizes pulsed infrared lasers as a source of stimulating radiation. The subject perceives such a stimulus as a color one due to the two-photon vision phenomenon [1].

The applicability of two-photon microperimetry depends largely on the parameters of the laser used for experiments. Therefore, in ICTER, we conduct extensive research on the influence of parameters of pulsed infrared laser, like pulse duration, pulse repetition rate, wavelength, on the perception by humans [2, 3]. Moreover, we perform a clinical assessment of two-photon microperimetry usefulness for earlier and more effective eye visual function abnormalities [4]. We hope that a deeper understanding of the phenomenon of two-photon vision, optimization of visual field test procedures, and clinical tests enable us to provide a useful tool for ophthalmologists worldwide. 

[1] Ruminski et al., BOE 10(9), pp. 4551-4567 (2019). DOI: 10.1364/BOE.10.004551

[2] Marzejon et al., BOE 12(2), pp. 462-479 (2021). DOI: 10.1364/BOE.411168

[3] Marzejon et al., Proc. SPIE 11623, 116231N (2021). DOI: 10.1117/12.2582735

[4] Komar et al., AOVS 62(8), 2009 (2021).

Text: Marcin Marzejon, MSc


Two-photon microperimetry: sensitivity of human photoreceptors to infrared light

Daniel Ruminski, Grazyna Palczewska, Maciej Nowakowski, Agnieszka Zielińska, Vladimir J. Kefalov, Katarzyna Komar, Krzysztof Palczewski, and Maciej Wojtkowski


Microperimetry is a subjective ophthalmologic test used to assess retinal function at various specific and focal locations of the visual field. Historically, visible light has been described as ranging from 400 to 720 nm. However, we previously demonstrated that infra-red light can initiate visual transduction in rod photoreceptors by a mechanism of two-photon absorption by visual pigments. Here we introduce a newly designed and constructed two-photon microperimeter. We provide for the first time evidence of the presence of a nonlinear process occurring in the human retina based on psychophysical tests using newly developed instrumentation. Since infra-red light penetrates the aged front of the eye better than visible light, it has the potential for improved functional diagnostics in patients with age-related visual disorders.

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ICTER scientists re-engineer the study of the cornea

A press release related to the paper “Multimode fiber enables control of spatial coherence in Fourier-domain full-field optical coherence tomography for in vivo corneal imaging” by Egidijus Auksorius, Dawid Borycki, and Maciej Wojtkowski, has been published on AlphaGalileo on July 29th, 2021.

Here is the AlphaGalileo press release:

A pioneer eye imaging discovery: ICTER scientists re-engineer the study of the cornea.


Telehealth applications

Smartphone-based optical palpation: towards elastography of skin for telehealth applications

Rowan W. Sanderson, Qi Fang, Andrea Curatolo, Aiden Taba, Helen M. DeJong, Fiona M. Wood, and Brendan F. Kennedy


Smartphones are now integral to many telehealth services that provide remote patients with an improved diagnostic standard of care. The ongoing management of burn wounds and scars is one area in which telehealth has been adopted, using video and photography to assess the repair process over time. However, a current limitation is the inability to evaluate scar stiffness objectively and repeatedly: an essential measurement for classifying the degree of inflammation and fibrosis. Optical elastography detects mechanical contrast on a micrometer- to millimeter-scale, however, typically requires expensive optics and bulky imaging systems, making it prohibitive for wide-spread adoption in telehealth. More recently, a new variant of optical elastography, camera-based optical palpation, has demonstrated the capability to perform elastography at low cost using a standard digital camera. In this paper, we propose smartphone-based optical palpation, adapting camera-based optical palpation by utilizing a commercially available smartphone camera to provide sub-millimeter resolution imaging of mechanical contrast in scar tissue in a form factor that is amenable to telehealth. We first validate this technique on a silicone phantom containing a 5 × 5 × 1 mm3 embedded inclusion, demonstrating comparative image quality between mounted and handheld implementations. We then demonstrate preliminary in vivo smartphone-based optical palpation by imaging a region of healthy skin and two scars on a burns patient, showing clear mechanical contrast between regions of scar tissue and healthy tissue. This study represents the first implementation of elastography on a smartphone device, extending the potential application of elastography to telehealth.

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Dr. Andrzej Foik – leader of the Ophthalmic Biology Group got granting under SONATA 16 funded by NSC

Source: www.ncn.gov.pl

On May 20, 2021, the Polish National Science Center announced the results of the SONATA 16 competition. One of the winners was our colleague, OBi group leader, Dr. Andrzej Foik. The topic of the winning competition is the role of the basal part of the crescentic brain in visual information processing. Find more information about the winning Project.


NSC announced the results of the competition for Polish-Lithuanian research projects – DAINA 2. Prof. Wojtkowski and Dr. Auksorius are among the winners

Source: www.ncn.gov.pl

On May 4, 2021 Polish National Science Center announced the results of the competition for Polish-Lithuanian research projects – DAINA 2. Prof. Wojtkowski and Dr. Auksorius are among the winners The topic of their winning project is Volumetric image reconstruction with filtering of redundant phase information. Find more information about the winning Project.


In vivo corneal imaging

Multimode fiber enables control of spatial coherence in Fourier-domain full-field optical coherence tomography for in vivo corneal imaging

Egidijus Auksorius, Dawid Borycki, and Maciej Wojtkowski


Fourier-domain full-field optical coherence tomography (FD-FF-OCT) has recently emerged as a fast alternative to point-scanning confocal OCT in eye imaging. However, when imaging the cornea with FD-FF-OCT, a spatially coherent laser can focus down on the retina to a spot that exceeds the maximum permissible exposure level. Here we demonstrate that a long multimode fiber with a small core can be used to reduce the spatial coherence of the laser and, thus, enable ultrafast in vivo volumetric imaging of the human cornea without causing risk to the retina.

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Two-photon vision phenomenon

Effects of laser pulse duration in two-photon vision threshold measurements

Marcin Marzejon, Łukasz Kornaszewski, Maciej Wojtkowski, Katarzyna Komar


Pulsed near-infrared (NIR) light sources can be successfully applied for both imaging and functional testing of the human eye, as published recently. These two groups of applications have different requirements. For imaging applications, the most preferable is invisible scanning beam while efficiently visible stimulating beam is preferable for functional testing applications. The functional testing of human eye using NIR laser beams is possible due to two-photon vision (2PV) phenomenon. 2PV enables perception of pulsed near-infrared laser light as color corresponding to approximately half of the laser wavelength. This study aims to characterize two-photon vision thresholds for various pulse lengths from a solidstate sub-picosecond laser (λc = 1043.3 nm, Frep = 62.65 MHz), either of 253 fs duration or elongated by Martinez- type stretcher to 2 ps, and fiber-optic picosecond laser (λc = 1028.4 nm, Frep = 19.19 MHz, τp = 12.2 ps).

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Adenine base editing

Retinal pigment epithelium (RPE) located at the back of the eye is essential for vision. It supports the photoreceptors, providing molecules required for their function. One of the main proteins produced by the RPE and indispensable for vision is the RPE65 enzyme, which is responsible for chemical signaling at the initial step of visual processing. De novo nonsense mutations in the Rpe65 gene underlie inherited genetic disorders of the eyes, resulting in blindness. To address this problem, we have harnessed the power of adenine base editors (ABEs) with Cas9 – single-guide RNA machinery to target  the mutations in the Rpe65 gene for their repair. We delivered genes coding for ABEs and the Cas9 system subretinally via a lentiviral vector. Our therapeutic manipulation corrected the pathogenic mutation in a mouse model with up to 29% efficiency and with minimal formation of indel and off-target mutations. The ABE-treated mice displayed restored RPE65 expression and its activity in the visual cycle. Moreover, we have observed near-normal levels of retinal and visual functions. Our findings motivate the further testing of ABEs for the treatment of inherited retinal diseases and for the correction of pathological mutations with non-canonical protospacer-adjacent motifs.


dr Andrzej Foik, e-mail: afoik@ichf.edu.pl & dr Anna Posłuszny, e-mail: aposluszny@ichf.edu.pl

Pertinent published article:

Restoration of visual function in adult mice with an inherited retinal disease via adenine base editing

Susie Suh, Elliot H. Choi, Henri Leinonen, Andrzej T. Foik, Gregory A. Newby, Wei-Hsi Yeh, Zhiqian Dong, Philip D. Kiser, David C. Lyon, David R. Liu & Krzysztof Palczewski, Nat Biomed Eng. 2021 Feb;5(2):169-178.