Details of a Researcher - Tsukada, Kosuke

Papers - Tsukada, Kosuke

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pH-taxis drives aerobic bacteria in duodenum to migrate into the pancreas with tumors.

Hiroaki Shirai, Cocoro Ito, Kosuke Tsukada

Scientific reports (Scientific Reports) 12 ( 1 ) 1783 - 1783 2022.12

ISSN 2045-2322

As oral or intestinal bacteria have been found in pancreatic cystic fluid and tumors, understanding bacterial migration from the duodenum into the pancreas via hepato-pancreatic duct is critical. Mathematical models of migration of aerobic bacteria from the duodenum to the pancreas with tumors were developed. Additionally, the bacterial distributions under the pH gradient and those under flow were measured in double-layer flow based microfluidic device and T-shaped cylinders. Migration of aerobic bacteria from the duodenum into pancreas is counteracted by bile and pancreatic juice flow but facilitated by pH-taxis from acidic duodenum fluid toward more favorable slightly alkaline pH in pancreatic juice. Additionally, the reduced flow velocity in cancer patients, due to compressed pancreatic duct by solid tumor, facilitates migration. Moreover, measured distribution of GFP E. coli under the pH gradient in a microfluidic device validated pH-tactic behaviors. Furthermore, Pseudomonas fluorescens in hydrochloride solution, but not in bicarbonate solution, migrated upstream against bicarbonate flow of > 20 μm/s, with an advancement at approximately 50 μm/s.

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Ex vivo sensing of primordial follicles in ovarian tissues by spectral-domain optical coherence tomography

Masaki Hosoda, Daisuke Oida, Koichiro Ito, Seido Takae, Nao Suzuki, Kosuke Tsukada

Medical Imaging 2022: Biomedical Applications in Molecular, Structural, and Functional Imaging (SPIE) 12036 2022

ISSN 0277-786X

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Convolutional neural network-based automatic detection of follicle cells in ovarian tissue using optical coherence tomography

Kasumi Saito, Yuki Motani, Seido Takae, Nao Suzuki, Kosuke Tsukada

Biomedical Physics & Engineering Express ({IOP} Publishing) 6 ( 6 ) 065026 - 065026 2020.09

Research paper (scientific journal), Joint Work, ISSN 2057-1976

To preserve the fertility of young female cancer patients, ovarian tissue cryopreservation and transplantation have been investigated as next-generation reproductive medical technologies. Noninvasive visualization of follicles in ovarian tissue and cryopreservation of higher density tissue is essential for effective transplantation. We proposed the use of optical coherence tomography (OCT) that can noninvasively visualize the internal structure of the ovarian tissue. However, a method for quantifying cell density has not yet been established because of the lack of available techniques to visualize follicles noninvasively. We proposed the use of a convolutional neural network (CNN) to extract small features from medical images as an image analysis method to automatically detect follicles from the obtained OCT images. First, we collected a total of 13 ovarian tissues from four-day-old mice and acquired OCT images using a full-field-type OCT. Then, the acquired images were analyzed using three detection methods: filter processing, filter processing combined with the CNN, and only CNN. Finally, to verify the detection accuracy of each method, the detection rate and precision were calculated by taking the doctor's detection as the correct result. The results showed that the detection method only using CNN achieved a detection rate of 0.81 and precision of 0.67; this indicated that follicles could be effectively detected using our proposed method. Furthermore, it is quantitatively evident that the density of follicles from the surface layer to the deep region differs depending on the tissue. In the future, these results could be used to detect follicles in tissues of different maturation stages and quantify follicles three-dimensionally, further accelerating next-generation reproductive medicine.

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Bacterial proteolytic activity improves drug delivery in tumors in a size, pharmacokinetic, and binding affinity dependent manner – A mechanistic understanding

Shirai H, Tsukada K

Journal of Controlled Release (Journal of Controlled Release) 321 348 - 362 2020.05

Joint Work, ISSN 01683659

© 2020 Elsevier B.V. Motile bacteria are able to penetrate in the distal areas of blood vessel, which makes bacteria attractive to researchers as a drug delivery vehicle carrying anti-cancer drugs to tumors. Not only therapeutic bacteria show wide anti-tumor effect but also the combination of therapeutic bacteria and conventional chemotherapy leads to dramatically large synergetic effect. We provide a mechanistic understanding of enhanced drug delivery in tumors by co-administration of chemotherapeutic agents and therapeutic bacteria. In this work, simultaneous delivery of C. novyi-NT and chemotherapeutic agents in tumors is mathematically modeled. Simulated doxorubicin concentration in tumors after Doxil administration with or without bacteria agreed reasonably well with experimental literature. Simulated doxorubicin concentration in tumors by the combination of Doxil and C. novyi-NT is over twice higher than that of Doxil alone. This enhanced doxorubicin concentration in tumors is due to the degradation of extracellular matrix of collagen by bacterial proteolytic activity, which increases hydraulic conductivity of interstitium, reduces interstitial fluid pressure, and thus increases convection through vessel walls. Additionally, it alleviates solid stress, which decompresses blood vessels, and thus increases vessel density. On the other hand, simulated doxorubicin concentration in tumors for non-liposomal free-doxorubicin is not enhanced by C. novyi-NT because vascular permeability of free-doxorubicin is larger than Doxil, and thus increased but relatively small convection across vessel walls is offset by the efflux due to increased interstitial flow. A strategy to further enhance this combination therapy is discussed along with sensitivity analysis.

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High-throughput single-cell live imaging of photobiomodulation with multispectral near-infrared lasers in cultured T cells

Katagiri W., Lee G.H., Tanushi A., Tsukada K., Choi H.S., Kashiwagi S.

Journal of Biomedical Optics (Journal of Biomedical Optics) 25 ( 3 ) 1 - 18 2020.03

ISSN 10833668

© 2020 by ASME. Significance: Photobiomodulation is a well-established therapeutic modality. However, the mechanism of action is poorly understood, due to lack of research in the causal relationship between the near-infrared (NIR) light irradiation and its specific biological effects, hindering broader applications of this technology. Aim: Since biological chromophores typically show several absorption peaks, we determined whether specific effects of photobiomodulation are induced with a combination of two wavelengths at a certain range of irradiance only, rather than a single wavelength of NIR light. Approach: In order to analyze a wide array of combinations of multispectral NIR light at various irradiances efficiently, we developed a new optical platform equipped with two distinct wavelengths of NIR lasers by high-throughput multiple dosing for single-cell live imaging. Two wavelengths of 1064 and 1270 nm were selected based on their photobiomodulatory effects reported in the literature. Results: A specific combination of wavelengths at low irradiances (250 to 400 mW/cm2 for 1064 nm and 55 to 65 mW/cm2 for 1270 nm) modulates mitochondrial retrograde signaling, including intracellular calcium and reactive oxygen species in T cells. The time-dependent density functional theory computation of binding of nitric oxide (NO) to cytochrome c oxidase indicates that the illumination with NIR light could result in the NO release, which might be involved in these changes. Conclusions: This optical platform is a powerful tool to study causal relationship between a specific parameter of NIR light and its biological effects. Such a platform is useful for a further mechanistic study on not only photobiomodulation but also other modalities in photomedicine.

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Respiration and Heat Shock Protein After Short-Term Heating/Stretch-Fixing on Smooth Muscle Cells

Kaminota N., Ogawa E., Kumagai H., Tsukada K., Arai T.

Cardiovascular Engineering and Technology (Cardiovascular Engineering and Technology) 2020

ISSN 1869408X

© 2020, Biomedical Engineering Society. Purpose: A treatment device without a stent is needed for peripheral stenotic artery treatment. We have proposed short-term heating balloon angioplasty, photo-thermo dynamic balloon angioplasty (PTDBA). Though smooth muscle cells (SMCs) after PTDBA are fixed in a stretched formation in a porcine model, influences of this stimulus on SMCs have not been investigated. SMC migration after vascular dilatation would be related to chronic restenosis. The aim of this study was to examine respiratory activity and recovery ability of SMCs after short-term heating/stretch-fixing in vitro for chronic phase treatment effect discussion. Methods: SMCs on a stretch chamber were heated for 15 s with stretching and fixed in a stretched formation. SMC migration is correlated with the cell respiratory activity. The amount of ATP production was measured using a WST-8 assay for respiratory activity evaluation. The intracellular expression of heat shock protein 70 was measured by an ELISA for recovery ability evaluation. Results: In the case of 60 °C heating, SMC respiratory activity after short-term heating/stretch-fixing decreased drastically in all stretching rates. In the case of 50 °C heating, SMC respiratory activity decreased and then increased. Alternatively, the recovery ability at 60 °C was greater than that at 50 °C. Conclusions: SMCs heated at 60 °C with stretching would have high recovery ability and low respiratory activity related to SMC migration. These results may be important evidence in determining the treatment condition in PTDBA.

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Development of a Patch-Type Flexible Oxygen Partial Pressure Sensor

Yuta Katayama, Yuta Fujioka, Kosuke Tsukada

IEEE Journal of Translational Engineering in Health and Medicine (Institute of Electrical and Electronics Engineers ({IEEE})) 8 1400607 - 7 2020

Research paper (scientific journal), Joint Work, ISSN 2168-2372

Oxygen concentration in living organisms is one of the important vital indicators in emergency care and bedside medical settings. However, the oximetry method has limitations: the measurement site is limited to the tissue containing blood and the absolute value of oxygen concentration cannot be measured. To overcome these limitations, in this work, we develop a new oxygen sensor that can directly measure the oxygen particle pressure ( $p\text{O}_{2}$ ) on the surface of the body and organs. A light emitting diode (LED) and a photodiode (PD) were embedded in a dimethylpolysiloxane substrate mixed with carbon nanotubes. The effectiveness of the device was evaluated using calibration, bending strain tests, time and frequency response, and finally in vivo assessments. The results reveal that the calibration experiment of the fabricated oxygen sensor device showed high sensitivity. The carbon nanotube electrode has a sufficient bending resistance and does not affect the response characteristics of the LED and PD, that is, it does not affect the oxygen measurement. In vivo assessment shows that the developed patch-type flexible oxygen sensor can accurately measure $p\text{O}_{2}$ by attaching it to tissues or organs having irregularities or curved surfaces and actual measurements on rat liver surface demonstrated its feasibility.

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Increasing surface-enhanced Raman scattering density using gold-coated magnetic nanoparticles controlled via a magnetic field for sensitive and efficient biomarker detection

Shibusawa K., Hase T., Tsukada K.

AIP Advances (AIP Advances) 9 ( 6 ) 2019.06

ISSN 2158-3226

© 2019 Author(s). Early detection of various diseases is expected using surface-enhanced Raman scattering (SERS). For example, a method of labeling an antibody of a disease-related molecule on metal nanoparticles and detecting the SERS signals of the particles bound to the antigen is a promising approach. However, the problems of a slow antigen-antibody reaction and low sensitivity remain unsolved. In this study, we fabricated nanoparticles that can be freely moved using an external magnetic field for rapid antigen-antibody reaction and also nanoengineered the substrate to increase the density of hotspots required for SERS. Gold-coated magnetic nanoparticles (Au-MNPs) with a core-shell structure were prepared by applying multiple coatings of gold onto magnetic iron(II,III) oxide nanoparticles, which were used as the core. A neodymium magnet easily moved and converged the Au-MNPs in the solution within a few seconds. In addition, a silver nanoparticle substrate (Ag-NS) with a hexagonal close-packed structure fixed on a polydimethylsiloxane thin film was prepared, and the stable generation of SERS was confirmed over the entire substrate. Upon aggregation of the Au-MNPs onto Ag-NS using a neodymium magnet, the total SERS strength per unit area drastically increased, suggesting that the combination of Au-MNPs and Ag-NS increased the density of the generated hotspots. In future work, with the labeling of antibodies onto Au-MNPs, we expect the proposed method to be applied in the sensitive measurement of biomarkers associated with diseases.

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Real-Time Imaging of Vaccine Biodistribution Using Zwitterionic NIR Nanoparticles

Katagiri W., Lee J.H., Tétrault M.A., Kang H., Jeong S., Evans C.L., Yokomizo S., Santos S., Jones C., Hu S., Fakhri G.E., Tsukada K., Choi H.S., Kashiwagi S.

Advanced Healthcare Materials (Advanced Healthcare Materials) 8 ( 15 ) e1900035 2019

ISSN 21922640

© 2019 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim Efficient and timely delivery of vaccine antigens to the secondary lymphoid tissue is crucial to induce protective immune responses by vaccination. However, determining the longitudinal biodistribution of injected vaccines in the body has been a challenge. Here, the near-infrared (NIR) fluorescence imaging is reported that can efficiently enable the trafficking and biodistribution of vaccines in real time. Zwitterionic NIR fluorophores are conjugated on the surface of model vaccines and tracked the fate of bioconjugated vaccines after intradermal administration. Using an NIR fluorescence imaging system, it is possible to obtain time-course imaging of vaccine trafficking through the lymphatics, observing notable uptake in lymph nodes with minimal nonspecific tissue interactions. Flow cytometry analysis confirmed that the uptake in lymph nodes by antigen presenting cells was highly dependent on the hydrodynamic diameter of vaccines. These results demonstrate that the combination of a real-time NIR fluorescence imaging system and zwitterionic fluorophores is a powerful tool to determine the fate of vaccine antigens. Since such non-specific vaccine uptake causes serious adverse reactions, this method is not only useful for optimization of vaccine design, but also for safety evaluation of clinical vaccine candidates.

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Micro/nano particle-based oxygen sensing film for monitoring respiration of cells cultured in a microfluidic device

Yabuki Y., Iwamoto Y., Tsukada K.

Japanese Journal of Applied Physics (Japanese Journal of Applied Physics) 58 ( SD ) 2019

ISSN 00214922

© 2019 The Japan Society of Applied Physics. Oxygen molecules are essential for producing energy in living organisms, meaning that oxygen measurement is required not only in the medical field, but also in research experiments. Porphyrin dyes in polystyrene microparticles were loaded and embedded in dimethylpolysiloxane to create oxygen sensing film. The lifetime of phosphorescence emitted from the film by laser excitation varied depending on the oxygen concentration, and the oxygen partial pressure was quantified from the lifetime. We revealed that the smaller the microparticle size and film thickness, the shorter the response time to the change in oxygen concentration. Next, the film was placed in cell culture microfluidic devices that can form oxygen gradients in the culture layer, and changes of the gradients due to oxygen metabolism of cells and implanted tumor tissues on the sensing film. From this, we demonstrated the effectiveness of the developed oxygen sensing film for medical and biological applications.

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金コート磁性ナノ粒子における表面増強ラマン散乱の発現と生体分子の検出

Fujino Masashi

Transactions of Japanese Society for Medical and Biological Engineering (Japanese Society for Medical and Biological Engineering) Annual57 ( Abstract ) S252_1 - S252_1 2019

ISSN 1347443X

ラマン分光法は生体分子や細胞を非侵襲的に検出する手法として期待されている．しかし，その散乱光強度は微弱であるため，表面増強ラマン散乱 (Surface Enhanced Raman Scattering: SERS) などによる増強機構を利用することで高感度化される．本研究では，SERS増強場の自在な制御と目的分子の高感度検出を目的として，磁性ナノ粒子に金をコートしたSERS発現用のナノ粒子を作製した． Fe3O4@Au (Micromod) にHAuCl4 とNH2OH・HClを攪拌しながら加える操作を繰り返すことで磁性ナノ粒子に金を複数回コートした．金のコート回数に伴う金と鉄の平均質量比の増加をEDX (Energy Dispersive X-ray spectrometry) 分析から定量した．ラマンマーカーとして4-ATP (4-Aminothiophenol) を金コート磁性ナノ粒子に修飾し，Renishaw製のin Via Raman microscopeを用いて4-ATPのラマンスペクトルを測定した．4-ATPに特徴的な1080cm-1 における信号強度がコート回数によって増減することを明らかにし，また最大強度を得るコート回数を決定した．金コート磁性ナノ粒子は外部磁場により液体中での移動を制御することが可能であり，将来的にはSERS増強場の自在な形成や生体分子の高感度化への応用が期待される．

Brief exposure of skin to near-infrared laser modulates mast cell function and augments the immune response

Kimizuka Y, Katagiri W, Locascio J, Shigeta A, Sasaki Y, Shibata M, Morse K, Sirbulescu R, Miyatake M, Reeves P, Suematsu M, Gelfand J, Brauns T, Poznansky M, Tsukada K, Kashiwagi S

Journal of Immunology (Journal of Immunology) 201 ( 12 ) 3587 - 3603 2018.12

Research paper (scientific journal), Joint Work, ISSN 00221767

©2018 by The American Association of Immunologists,Inc. The treatment of skin with a low-power continuous-wave (CW) near-infrared (NIR) laser prior to vaccination is an emerging strategy to augment the immune response to intradermal vaccine, potentially substituting for chemical adjuvant, which has been linked to adverse effects of vaccines. This approach proved to be low cost, simple, small, and readily translatable compared with the previously explored pulsed-wave medical lasers. However, little is known on the mode of laser-tissue interaction eliciting the adjuvant effect. In this study, we sought to identify the pathways leading to the immunological events by examining the alteration of responses resulting from genetic ablation of innate subsets including mast cells and specific dendritic cell populations in an established model of intradermal vaccination and analyzing functional changes of skin microcirculation upon the CW NIR laser treatment in mice. We found that a CW NIR laser transiently stimulates mast cells via generation of reactive oxygen species, establishes an immunostimulatory milieu in the exposed tissue, and provides migration cues for dermal CD103+ dendritic cells without inducing prolonged inflammation, ultimately augmenting the adaptive immune response. These results indicate that use of an NIR laser with distinct wavelength and power is a safe and effective tool to reproducibly modulate innate programs in skin. These mechanistic findings would accelerate the clinical translation of this technology and warrant further explorations into the broader application of NIR lasers to the treatment of immune-related skin diseases. The Journ Al of Immunology, 2018, 201: 3587-3603.

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Preliminary human application of optical coherence tomography for quantification and localization of primordial follicles aimed at effective ovarian tissue transplantation

Takae S., Tsukada K., Maeda I., Okamoto N., Sato Y., Kondo H., Shinya K., Motani Y., Suzuki N.

Journal of Assisted Reproduction and Genetics (Journal of Assisted Reproduction and Genetics) 35 ( 4 ) 627 - 636 2018.04

ISSN 10580468

© 2018, The Author(s). Purpose: The purpose of this study was to evaluate the possible clinical application of optical coherence tomography for assessing ovarian reserve in individual specimens of human ovarian tissue for fertility preservation. Methods: Ovarian tissue examination by optical coherence tomography was performed before ovarian tissue cryopreservation. Three of the four subjects had hematological disease or cancer, and they faced a threat to their fertility due to impending chemotherapy. One patient underwent ovarian tissue extraction for in vitro activation of dormant follicles as fertility treatment. Results: The current full-field optical coherence tomography technique can detect primordial follicles in non-fixed and non-embedded human ovarian tissue. These images are well correlated with histological evaluation and the ovarian reserve test, including follicle counts. Conclusion: It was demonstrated that optical coherence tomography could assess localization of primordial follicles and ovarian reserve in specimens of non-fixed human ovarian cortex, although optimization for examination of human ovarian tissue is needed for clinical application. Additionally, this technique holds the possibility of assessing the ovarian reserve of patients with unevaluable ovarian reserve. Trial registration number: UMIN000023141.

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Immobilized monolayer nanoparticles in a microfluidic device for surface enhanced Raman scattering measurement

Hase T., Ishigaki S., Shibusawa K., Hamanaka S., Yabuki Y., Tamano Y., Tsukada K.

Advanced Biomedical Engineering 6 122 - 128 2018.01

Research paper (scientific journal), Joint Work, Accepted

Development of bioadhesive oxygen sensor using electrochemiluminescence

KATAYAMA YUTA, FUJIOKA YUTA, TSUKADA KOSUKE

Transactions of Japanese Society for Medical and Biological Engineering (Japanese Society for Medical and Biological Engineering) Annual56 ( Abstract ) S88 - S88 2018

ISSN 1347443X

Noninvasive continuous oxygen measurement in living bodies is important in clinics and basic experiments. Focusing on the phosphorescent-lifetime method which can measure the absolute value of the oxygen concentration, we developed a film type oxygen sensor. A thin oxygen sensing film was prepared by mixing Pd-meso-tetra(4-carboxyphenyl)porphyrin (Pd-TCPP) with Polydimethylsiloxane (PDMS). We report calibration experiments on oxygen partial pressure change by using electrochemiluminescenece (ECL) and LED (Light emitting diode) as light source and electrode characteristics for flexible substrate material. AC voltage was applied to the light source, and the oxygen partial pressure was calculated from phase shift of the light source and phosphorescence. As a result of calibration experiment, we could measure oxygen-partial-pressure with high accuracy by LED. However, ECL excitation showed a phase shift, but measurement accuracy was lower than LED. We will improve the measurement accuracy by improving emission intensity and applied voltage pattern of ECL.

近赤外半導体レーザーデバイスによる免疫活性化作用の検証

片桐渉, 君塚善文, Kashiwagi Satoshi, 塚田孝祐

日本レーザー医学会誌 ((NPO)日本レーザー医学会) 38 ( 3 ) 287 - 287 2017.10

ISSN 0288-6200

Biodegradable polycaprolactone nanofibres with β-chitosan and calcium carbonate produce a hemostatic effect

Park J.Y., Kyung K.H., Tsukada K., Kim S.H., Shiratori S.

Polymer (ELSEVIER) 123 194 - 202 2017.08

Research paper (scientific journal), Joint Work, Accepted, ISSN 0032-3861

We newly developed high-performance blood coagulation nanofibres using Polycaprolactone (PCL) including Calcium carbonate (CaCO3) and beta-chitosan. The most important feature for a wound dressing is that it is harmless to the human body. Here, we fabricated the nanofibre using all human-safe materials. We used a PCL nanofibre mat as a substrate and then synthesized artificial CaCO3 from sodium carbonate and calcium chloride. The CaCO3 was then added to the fibre solution to create PCL/CaCO3 nanofibres. We coated PCL and PCL/CaCO3 nanofibres with b-chitosan as a hemostatic material via a spray method. For the more uniform coating, we used ultrasonic spray coating method and then compared the blood coagulation abilities of the PCL and PCL/CaCO3 nanofibres. We found that the PCL/CaCO3 nanofibres sprayed with beta-chitosan had a greater effect on blood coagulation than the PCL nanofibres. In the result of animal experiment, beta-chitosan have a key role in changing of surface wettability from hydrophobic to hydrophilic. Moreover, this is contributed to enhance blood coagulability. PCL/CaCO3 nanofibres sprayed with beta-chitosan therefore offer promise in medical applications. (C) 2017 Elsevier Ltd. All rights reserved.

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Near-infrared 1064 nm laser modulates migratory dendritic cells to adjuvant the immune response to intradermal influenza vaccine

Morse K., Kimizuka Y., Chan M.P.K., Shibata M., Shimaoka Y., Takeuchi S., Forbes B., Nirschl C.J., Li B., Zeng Y., Bronson R., Katagiri W., Shigeta A., Sirbulescu R.F., Chen H., Tan R.Y.Y., Tsukada K., Brauns T., Gelfand J.A., Sluder A., Locascio J.J., Poznansky M.C., Anandasabapathy N., Kashiwagi S.

The Journal of Immunology (The American Association of Immunologists) 199 ( 4 ) 1319 - 1332 2017.07

Research paper (scientific journal), Joint Work, Accepted

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Semiconductor diode laser device adjuvanting intradermal vaccine

Yoshifumi Kimizuka, John J. Callahan, Zilong Huang, Kaitlyn Morse, Wataru Katagiri, Ayako Shigeta, Roderick Bronson, Shu Takeuchi, Yusuke Shimaoka, Megan P.K. Chan, Yang Zeng, Binghao Li, Huabiao Chen, Rhea Y.Y. Tan, Conor Dwyer, Tyler Mulley, Pierre Leblanc, Calum Goudie, Jeffrey Gelfand, Kosuke Tsukada, Timothy Brauns, Mark C. Poznansky, David Bean, Satoshi Kashiwagi

Vaccine (Elsevier) 35 ( 18 ) 2404 - 2412 2017.04

Research paper (scientific journal), Joint Work, Accepted

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Accuracy and safety verification of ovarian reserve assessment technique for ovarian tissue transplantation using optical coherence tomography in mice ovary

Seido Takae, Kosuke Tsukada, Yorino Sato, Naoki Okamoto, Tai Kawahara, Nao Suzuki

Scientific Reports (Nature Publishing Group) 7 43550 2017.03

Research paper (scientific journal), Joint Work, Accepted, ISSN 2045-2322

Except for histological study, there are currently no suitable techniques available for the detection and identification of primordial follicles in ovary of primary ovarian insufficiency patients who have undetectable AMH levels. Also, the ability to locate and quantify follicles on ovarian cortex strips, without fixation, is valuable for patients who could undergo subsequent successful ovarian tissue transplantation. Although optical coherence tomography (OCT) is a well-established high resolution imaging technique without fixation commonly applied in biomedicine, few reports are available on ovarian tissue imaging. In present study, we established standard OCT follicle images at each developmental stage, including the primordial follicle, and demonstrated the efficacy of OCT to estimate IVF outcome in transplanted mice ovary like ovarian reserve tests. Unfortunately, the current commercial OCT could not be used to accurate follicle count the number of follicles for whole ovary, because the maximum depth of examination was 100 mu m. And we demonstrated the safety of OCT examination, it did not affect IVF outcome and birth defect rate, and reproductive ability. Although there is room for improvement, these findings will be first step to bring OCT examination a step closer to clinical application for measuring true ovarian reserve and localizing follicles.

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