Our ongoing projects include a coherent optical adaptive technique-based two-photon microscopy, acoustic wave guided optical phase conjugation, and ultra-high speed two-photon microscopy for pushing the speed of deep tissue fluorescence imaging to the theoretical limit. Moreover, several other new techniques are being designed, including multiplex stimulated emission microscopy, multichannel excitation two-photon microscopy, and novel one-photon microscopy for deep tissue fluorescence imaging. The ultimate goal is to advance the frontier of optical molecular imaging and sensing to provide unprecedented detection depth, quality, sensitivity, and speed for a broad range of applications in biology and medicine
Elastic scattering is the dominant process during optical wave propagation in tissues. If there is a way to suppress elastic scattering, the penetration depth of optical imaging could be improved by about two orders of magnitudes, which could open up many possibilities for both scientific research and medical applications.
The two most widely used accurate deep tissue optical imaging techniques are Optical Coherence Tomography (OCT) and two-photon fluorescence microscopy. OCT detects the ballistic light reflection and can provide ~1 mm penetration depth. Two-photon microscopy employs ballistic light for illumination and collects both scattered and ballistic fluorescence signals. Unlike OCT, two-photon microscopy provides molecular contrast (fluorescence) and the penetration depth is ~0.5 mm. Using longer excitation wavelength (1,200-1,300 nm), two-photon microscopy can achieve improved penetration depth (~1mm). At greater penetration depth, the ballistic component could be easily overwhelmed by the out-of-focus scattered components. In addition, the accumulated aberration becomes significant. Adaptive optics has been employed to compensate for the aberration in optical microscopy. To date, the successful demonstrations have shown that it is possible to improve the image quality (resolution, signal strength). However, the improvement in optical penetration depth is moderate. To further increase the penetration depth, novel methods that employ not only the ballistic light but also the scattered light for imaging should be explored. One of our primary research goals is to develop new technologies to control and use the scattered light for deep tissue molecular imaging.
Another important line of research is the development of hybrid imaging methods that combine light with sound. One method is to use light absorption to generate sound, a method called photoacoustic tomography. The other method is to use sound to modulate light, known as ultrasound modulated optical tomography. These hybrid methods, which combine the optical imaging contrast (absorption) with the penetration depth of the ultrasound, are very promising for clinical applications. However, the sensitivity and the spatial resolution are far from start-of-the-art optical microscopy. Our laboratory is developing a new type of hybrid imaging method that may potentially provide higher sensitivity and spatial resolution.
For practical biomedical applications, imaging speed is very important. High-speed imaging can allow scientists to capture very fast biological events. One of our research goals is to develop a robust and flexible ultra-high speed deep tissue imaging technology that can provide imaging speed at the theoretical limit (fluorescence decay rate) in deep tissues. Unlike previous attempts, our method can conveniently work with commercially available laser sources and the total data rate can be easily varied to match the decay rate of the fluorophore.
The Laser was first demonstrated at the Hughes Research Laboratory in 1960. Now, half a century has passed and the development of laser technologies continues to change both scientific research and our daily life. The capabilities of laser science for revealing the properties of materials and for performing measurement at unprecedented sensitivity and resolution are truly amazing. In addition to optical imaging, our laboratory is exploring new technologies of using laser to perform sensitive measurements.
Random scattering and aberrations severely limit the imaging depth in optical microscopy. We introduce a rapid, parallel wavefront compensation technique that efficiently compensates even highly complex phase distortions. Using coherence gated backscattered light as a feedback signal, we focus light deep inside highly scattering brain tissue. We demonstrate that the same wavefront optimization technique can also be used to compensate spectral phase distortions in ultrashort laser pulses using nonlinear iterative feedback. We can restore transform limited pulse durations at any selected target location and compensate for dispersion that has occurred in the optical train and within the sample.
Optical microscopy has so far been restricted to superficial layers, leaving many important biological questions unanswered. Random scattering causes the ballistic focus, which is conventionally used for image formation, to decay exponentially with depth. Optical imaging beyond the ballistic regime has been demonstrated by hybrid techniques that combine light with the deeper penetration capability of sound waves. Deep inside highly scattering media, the sound focus dimensions restrict the imaging resolutions. Here we show that by iteratively focusing light into an ultrasound focus via phase conjugation, we can fundamentally overcome this resolution barrier in deep tissues and at the same time increase the focus to background ratio. We demonstrate fluorescence microscopy beyond the ballistic regime of light with a threefold improved resolution and a fivefold increase in contrast. This development opens up practical high resolution fluorescence imaging in deep tissues.
Ultrasound pulse guided digital phase conjugation has emerged to realize fluorescence imaging inside random scattering media. Its major limitation is the slow imaging speed, as a new wavefront needs to be measured for each voxel. Therefore 3D or even 2D imaging can be time consuming. For practical applications on biological systems, we need to accelerate the imaging process by orders of magnitude. Here we propose and experimentally demonstrate a parallel wavefront measurement scheme towards such a goal. Multiple focused ultrasound pulses of different carrier frequencies can be simultaneously launched inside a scattering medium. Heterodyne interferometry is used to measure all of the wavefronts originating from every sound focus in parallel. We use these wavefronts in sequence to rapidly excite fluorescence at all the voxels defined by the focused ultrasound pulses. In this report, we employed a commercially available sound transducer to generate two sound foci in parallel, doubled the wavefront measurement speed, and reduced the mechanical scanning steps of the sound transducer to half.
Fluorescence imaging has revolutionized biomedical research over the past three decades. Its high molecular specificity and unrivalled single-molecule-level sensitivity have enabled breakthroughs in a number of research fields. For in vivo applications its major limitation is its superficial imaging depth, a result of random scattering in biological tissues causing exponential attenuation of the ballistic component of a light wave. Here, we present fluorescence imaging beyond the ballistic regime by combining single-cycle pulsed ultrasound modulation and digital optical phase conjugation. We demonstrate a near-isotropic three-dimensional localized sound–light interaction zone. With the exceptionally high optical gain provided by the digital optical phase conjugation system, we can deliver sufficient optical power to a focus inside highly scattering media for not only fluorescence imaging but also a variety of linear and nonlinear spectroscopy measurements. This technology paves the way for many important applications in both fundamental biology research and clinical studies.
Superpenetration optical microscopy by iterative multiphoton adaptive compensation technique.Proceedings of the National Academy of Sciences of the United States of America 2012
J. Tang, R. N. Germain, and M. Cui Proceedings of the National Academy of Sciences of the United States of America, 109:8434-9 (2012)
Biological tissues are rarely transparent, presenting major challenges for deep tissue optical microscopy. The achievable imaging depth is fundamentally limited by wavefront distortions caused by aberration and random scattering. Here, we report an iterative wavefront compensation technique that takes advantage of the nonlinearity of multiphoton signals to determine and compensate for these distortions and to focus light inside deep tissues. Different from conventional adaptive optics methods, this technique can rapidly measure highly complicated wavefront distortions encountered in deep tissue imaging and provide compensations for not only aberration but random scattering. The technique is tested with a variety of highly heterogeneous biological samples including mouse brain tissue, skull, and lymph nodes. We show that high quality three-dimensional imaging can be realized at depths beyond the reach of conventional multiphoton microscopy and adaptive optics methods, albeit over restricted distances for a given correction. Moreover, the required laser excitation power can be greatly reduced in deep tissues, deviating from the power requirement of ballistic light excitation and thus significantly reducing photo damage to the biological tissue.
We show through experiments and simulations that parallel phase modulation, a technique developed in the field of adaptive optics, can be employed to quickly determine the spectral phase profile of ultrafast laser pulses and to perform phase compensation as well as pulse shaping. Different from many existing ultrafast pulse measurement methods, the technique reported here requires no spectrum measurements of nonlinear signals. Instead, the power of nonlinear signals is used directly to quickly measure the spectral phase, a convenient feature for applications such as two-photon fluorescence microscopy. The method is found to work with both smooth and even completely random distortions. The experimental results are verified with MIIPS measurements.
A large number of degrees of freedom are required to produce a high quality focus through random scattering media. Previous demonstrations based on spatial phase modulations suffer from either a slow speed or a small number of degrees of freedom. In this work, a high speed wavefront determination technique based on spatial frequency domain wavefront modulations is proposed and experimentally demonstrated, which is capable of providing both a high operation speed and a large number of degrees of freedom. The technique was employed to focus light through a strongly scattering medium and the entire wavefront was determined in 400 milliseconds, ~three orders of magnitude faster than the previous report.
A parallel wavefront optimization method is demonstrated experimentally to focus light through random scattering media. The simultaneous modulation of multiple phase elements, each at a unique frequency, enables a parallel determination of the optimal wavefront. Compared to a pixel-by-pixel measurement, the reported parallel method uses the target signal in a highly efficient way. With 441 phase elements, a high-quality focus was formed through a glass diffuser with a peak-to-background ratio of ∼270. The accuracy and repeatability of the system were tested through experiments.
One postdoc position is immediately available for highly motivated and skilled experimentalists. Research background in optical science and engineering (such as optical microscopy, spectroscopy, AMO physics, nonlinear optics, holography, adaptive optics, laser optics, and ultrafast optics) is preferred. The ultimate goal of our lab is to develop new technologies that may impact the way biomedical research is performed. Strong interest towards this line of research is a plus. For application, please send email to Dr. Meng Cui.
We have multiple openings for visiting graduate students. Janelia will provide housing + $1000/month living expenses. These projects are for the development of next generation biophotonics technologies. The typical duration for one project is six months. Research background in physics, optics and electrical engineering are preferred. To apply, please send email to Dr. Meng Cui.