Bacteriorhodopsin as a high-resolution, high-capacity buffer for digital ...

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Mar 19, 2004 - holography has been successfully demonstrated in recent years, unfortunately the limited information capacity of present electronic sensors,.
INSTITUTE OF PHYSICS PUBLISHING

MEASUREMENT SCIENCE AND TECHNOLOGY

Meas. Sci. Technol. 15 (2004) 639–646

PII: S0957-0233(04)67659-4

Bacteriorhodopsin as a high-resolution, high-capacity buffer for digital holographic measurements D H Barnhart1, W D Koek2, T Juchem3, N Hampp3, J M Coupland4 and N A Halliwell4 1

Department of Theoretical and Applied Mechanics, University of Illinois at Urbana-Champaign, Urbana, IL 61801-2983, USA 2 Optics Research Group, Delft University of Technology, The Netherlands 3 Institute for Physical Chemistry, University of Marburg, Hans-Meerwein-Strasse Geb. H, D-35032 Marburg, Germany 4 Wolfson School of Mechanical and Manufacturing Engineering, Loughborough University, Loughborough, Leicestershire LE11 3TU, UK E-mail: [email protected]

Received 15 August 2003, in final form 22 January 2004 Published 19 March 2004 Online at stacks.iop.org/MST/15/639 (DOI: 10.1088/0957-0233/15/4/005) Abstract Recent trends in optical metrology suggest that, in order for holographic measurement to become a widespread tool, it must be based on methods that do not require physical development of the hologram. While digital holography has been successfully demonstrated in recent years, unfortunately the limited information capacity of present electronic sensors, such as CCD arrays, is still many orders of magnitude away from directly competing with the high-resolution silver halide plates used in traditional holography. As a result, present digital holographic methods with current electronic sensors cannot record object sizes larger than several hundred microns at high resolution. In this paper, the authors report on the use of bacteriorhodopsin (BR) for digital holography to overcome these limitations. In particular, BR is a real-time recording medium with an information capacity (5000 line-pairs/mm) that even exceeds high resolution photographic film. As such, a centimetre-square area of BR film has the same information capacity of several hundred state-of-the-art CCD cameras. For digital holography, BR temporarily holds the hologram record so that its information content can be digitized for numeric reconstruction. In addition, this paper examines the use of BR for optical reconstruction without chemical development. When correctly managed, it is found that BR is highly effective, in terms of both quality and process time, for three-dimensional holographic measurements. Consequently, several key holographic applications, based on BR, are proposed in this paper. Keywords: bacteriorhodopsin, holographic velocimetry, three-dimensional

measurements, digital holography

1. Introduction Recent trends in optical metrology suggest that, in order for holographic measurement to become a widespread tool, it must be based on methods that do not require physical 0957-0233/04/040639+08$30.00

development. In particular, traditional holograms recorded on photographic film require time-consuming chemical development procedures that inhibit its ease of use for many applications. In recent years, however, digital holographic recording and reconstruction (Schnars and

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J¨uptner 1994, Jacquot et al 2001, Sun et al 2002), digital holographic displacement measurement on micro-components (Seebacher et al 1997) and digital holographic velocity (HV) measurement in miniature flows (Skarman et al 1996, 1999, Co¨etmellec et al 2001) have all been successfully demonstrated. While such digital holographic methods are unparalleled in their simplicity and ease of use, unfortunately the limited information capacity of present electronic sensors, such as CCD arrays, is still orders of magnitude away from directly competing with high-resolution photographic film. As such, digital holograms that have been directly recorded on present-day CCD sensors cannot take comparable highresolution measurements from object volumes larger than several hundred microns in size. Otherwise, the higher volumetric measurement size must be traded for diminished measurement resolution. This paper introduces an approach that uses bacteriorhodopsin (BR) film as an intermediate information buffer to store the holographic information for further analysis. In particular, BR is a real-time recording medium with an information capacity (5000 line-pairs/mm) that even exceeds high-resolution photographic film. In many ways, BR appears well suited for digital holography. Holograms based on BR offer both high resolution and ease of use. In contrast to the current state-of-the-art camera sensor array, which may record more than 20 million pixels of information, a digital holographic camera based on BR can record 1–100 billion pixels of information. This corresponds to a holographic object space volume of 1–100 cm3 with sub-micron spatial resolution. In comparison, a BR hologram of similar dimensions to the CCD chip size can resolve the same object resolution from a million times greater volume. Although the development of solid-state camera sensor technology continues to improve at a rapid pace, it still appears that conventional camera sensors will not reach the information capacity of BR film for years to come. With digital holographic measurement, the BR film temporarily holds the hologram record so that its information content can be digitized for numeric analysis. BR can deliver a reconstruction signal-to-noise ratio of 50 dB as well as an object-light recording sensitivity of 50 µJ cm−2 that corresponds with Agfa 8E56 holographic emulsion (Juchem and Hampp 2001). Although BR still has an overall light sensitivity requirement of ∼1–5 mJ cm−2, only 50 µJ cm−2 needs to be present in the scattered object light and the remainder of the required energy can be placed in the reference beam. (Note, however, that this does not consider the initial energy requirements for object illumination: only the object-scattered light that reaches the hologram is being considered here.) In contrast with photorefractive crystals, which have restricted apertures, BR holograms can be produced in large formats (100 × 100 mm2) with large numeric apertures. Unlike thermoplastics, which can only support transmission holograms of restricted geometry, BR can be used in either reflection or transmission formats without restriction. Finally, unlike many real-time materials which have cycle-limited lifetimes, BR holograms can be recycled indefinitely without degradation. 640

2. Physical properties of BR for holography BR has a number of properties that are worth further consideration (Juchem and Hampp 2001). BR is a photochromic material that switches between purple and yellow colours with its main sensitivity in the blue and green portions of the optical spectrum. In many cases, BR holograms are recorded at a green (∼532 nm) laser wavelength and erased with a blue source of light using wavelengths between 400 and 450 nm. In general, it is not necessary for the erasing source to be coherent and, very often, a filtered flash lamp is used instead of a laser. However, the roles of the two wavelengths can also be reversed with a blue recording laser and a green erasure beam. In special cases, the blue and green CW lasers can be configured to operate simultaneously, where the green laser wavelength continuously reconstructs the holographic image while a blue laser is simultaneously recording the hologram (Hampp et al 1990). Even though the result is an actively pumped hologram whose image is dynamically refreshed, this novel configuration does not work with pulsed laser recording and therefore is not normally considered for particle holography. When reconstructed at the 532 nm wavelength, BR works as an absorption hologram and has a diffraction efficiency of about 1%. While BR is not very sensitive to red wavelengths, such as 632 nm, it operates as a phase grating at these longer wavelengths and can diffract light more efficiently by a factor of 2 or greater. Finally, BR has an interesting property in that it records and reconstructs the relative polarization of the object wave front. This property can be exploited to increase the signal-to-noise ratio of the reconstruction or to record two polarization-independent holograms on the same BR plate. This presents a powerful means to eliminate the effect of stray light reflections from optical windows present in the experiment. In particular, if the recorded object depolarizes the light while the specular background reflections remain polarized, the stray window light can be filtered out and the recorded object information can be recovered with a linear polarizer placed in front of the hologram during reconstruction. An experimental example of this phenomenon is presented below in figure 1. Here, a hologram was recorded of sub-micron aerosol particles located in front of an optical window. In this case, the illumination beam was directed to scatter backwards off both the particles and the optical window. Although the hologram recorded the window reflections as well as the particle images, the polarization of window reflections remained collinear with the illumination beam while the particle images were depolarized. Since the original polarization states were preserved in the BR hologram, the depolarized component of the particle signal (figure 1(b)) during holographic reconstruction could be recovered independently from the background specular noise (figure 1(a)). This example also demonstrates the high image resolution of BR-recorded holograms. Unlike many other holographic materials that exhibit effects from emulsion shrinkage (Barnhart et al 1994), BR holograms do not suffer from distortion in the recorded grating. As a consequence, the real holographic image resolution (as shown in figure 1(b)) can exceed the sampling resolution of the CCD camera (∼7 µm in this case).

Bacteriorhodopsin as a high-resolution, high-capacity buffer for digital holographic measurements

Figure 1. (a) Specular background noise from optical windows. (b) Optically filtered and digitally thresholded particle images in the same spatial position as (a).

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In many respects, bacteriorhodopsin (BR) appears to be ideally suited for holography. Nevertheless, the material does possess limitations that must be taken into account. As mentioned previously, the light sensitivity of BR for holography depends on having the correct reference beam energy. Unlike the inherent tolerance of silver halide holography to varying degrees of light exposure, BR holography demands careful controlling of the reference beam energy. In particular, the reference energy must be adjusted to optimize the total plate exposure. As such, the applied reference beam energy depends on the scattered energy received from the object. As shown by the lower arrow in figure 2, the optimal exposure biases the bleach curve to the linear region, where the slope is the steepest and the fringe modulation is the most effective. For this particular BR material, the reference energy needs to be adjusted to deliver a total exposure of 5 mJ cm−2 for CW lasers and 3 mJ cm−2 for Q-switched lasers. In instances when the beam ratio cannot be adequately controlled or when insufficient laser energy is available, the outcome may not be successful. As such, with dynamic measurements that require a pulsed laser, the available laser pulse energy can place an upper limit on the size of the recorded BR hologram as well as the maximum size of the potential object space. This may in turn have an effect on the type of application that can be effectively managed. Nevertheless, the

nature of object subject matter as well as the chosen hologram geometry can change these limitations dramatically. As such, it is difficult to place specific values on these limitations. However, a rule of thumb might be that between 100 mJ and 1000 mJ of pulsed laser energy can be effectively employed for many applications where dynamic processes are involved. This, in turn, permits hologram sizes that range from 1 to 100 cm2 and object-space volumes between 1 and 100 cm3. For still object measurements, however, a continuous laser can be employed with relaxed constraints concerning the overall hologram and object-space dimensions. Another limitation of BR holograms is the limited life of the recorded fringe information. In particular, at room temperature, a BR hologram image will thermally decay and become significantly degraded after only a few minutes. This limited time span may also restrict the amount of information that can be recovered from the hologram. Hence, the speed at which the hologram analysis takes place needs to be maximized in order to recover the greatest amount of information from the hologram. However, the available time span can be considerably increased by reducing the hologram temperature below room temperature. Although the specific gains will vary according to the BR formulation and optical geometry, for reduced temperatures of 0–5 ◦ C, the hologram image lifetime can be extended by a factor of ten and even greater gains are possible at lower temperatures.

3. Some generic digitization strategies While BR holograms can certainly be viewed by optical reconstruction alone, today’s easy availability of fast and inexpensive computers with hundreds of gigabytes of harddisk space finally makes numeric reconstruction an attractive alternative. However, before a computer can do anything, the hologram must first be digitized and the recorded wave front stored as a table of complex numbers. After this has been accomplished, a digital Fresnel-transform can be applied to the numeric complex-field data to reconstruct the object information at a particular image plane and numerically investigate the image space within the computer’s memory. There can be many different approaches for digital recording and numeric reconstruction with BR. However, all approaches involve taking a digital scan of fringe intensity 641

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Recording Reference Beam Object

Object Illumination Hologram

Figure 3. Recording of the hologram.

Incoherent Beam CCD Microscope

Hologram Figure 4. Incoherent sampling of the hologram.

information. This digitization process can use either coherent or incoherent sampling of the fringes. In both cases, this paper proposes to digitize the BR hologram by taking twodimensional samples of the recorded wave front at evenly spaced, overlapping intervals across the wave-front aperture. This can be accomplished either with a two-dimensional CCD camera that samples square patches of the fringe information or with a line-scan camera that samples stripes of information. After the initial samples are digitized, the patches of information are stitched together to create a seamless composite of the hologram information. Consider the initial holographic recording geometry shown in figure 3. We will next examine three different strategies to extract information from this hologram. The first strategy, shown in figure 4, uses an incoherentsampling method. Here, the hologram is illuminated with an incoherent light source and very small patches of the recorded fringes on the hologram surface are incoherently imaged into a video microscope at high magnification. The scanned patches are then stitched together to create a seamless image of the hologram grating surface. This technique permits the BR hologram to be scanned at a wavelength that is different from the original recorded wavelength, which retards the hologram erasure during the digital scanning process. In 642

particular, while BR holograms remain optically sensitive during the readout process, the sensitivity is greatly reduced at longer wavelengths. Unfortunately, incoherent imaging procedures are resolution-limited by the point-spread function of the microscope imaging system. Another limitation is the short depth-of-focus tolerances of such high-magnification imaging systems. As a result, the scanning process must be tightly controlled to maintain a constant microscope distance to the hologram surface throughout the scanning process. Finally, incoherent imaging can only be used with transmission holograms and not Bragg-reflection hologram geometries, since reflection hologram fringes cannot be incoherently imaged. The remaining two strategies rely on coherent-sampling methods. Shown below in figures 5(a) and (b), the hologram is now illuminated by a coherent light source and the hologram-diffracted wave front is then digitized by mixing the holographic reconstruction with a second reference beam at the camera sensor plane. In this case, the digitized information can either coincide with the hologram surface, as shown in figure 5(a), or it may be located at any chosen distance from the hologram, as shown in figure 5(b). With these two latter methods, the hologram information is first converted into complex-field information before it is stitched together. To accomplish this, the local mixing reference beam can either be placed off-axis to create carrier fringes or located in-line and phase-stepping methods employed to unwrap and recover the recorded phase information (Vest 1979). With the coherent sampling methods, an optical Fouriertransform is often employed in place of the imaging lens used with incoherent sampling methods. As such, spatial frequencies in the hologram space are mapped into spatial positions at the Fourier plane. This has the consequence that the size of the CCD pixels no longer limits the resolving power of the digitizing system. Instead, the spatial resolution of the CCD sensor limits the maximum size of the wave-front sampling aperture, but the system resolving power is solely determined by the numeric aperture of the Fourier-transform optics. In practice, the optical Fourier transform can be accomplished with either an optical Fourier-transform lens (and a planar reference beam), depicted in figure 5, or by a lens-less Fourier-transform method (and a spherical reference beam) (Goodman 1968). In either case, the optical Fourier-transform process removes the excess quadratic phase curvature from the object wave front which minimizes the maximum spatial frequency presented to the CCD array. In order to restrict the wave-front sampling aperture, two different methods are employed that depend on the camera geometry. The first method, shown in figure 5(a), limits the size of the reconstruction beam. This technique is possible when the wave front is sampled on the hologram surface. The second method, shown in figure 5(b), uses a physical pinhole aperture and is used when the wave-front sample plane is not located at the hologram plane. Finally, coherent-sampled holograms have a big advantage over incoherent methods in that the fringe sampling procedures are not resolution-limited by aberrations present in the camera optics. If the transfer-function of the lens system can be accurately characterized, then the effects of aberrations

Bacteriorhodopsin as a high-resolution, high-capacity buffer for digital holographic measurements

Coherent Digital Scan Local Reference

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Figure 5. (a) Coherent sampling of the hologram plane. (b) Coherent sampling of the image plane. Two Converging Reference Waves

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Figure 6. (a) Recording of the hologram. (b) Object-conjugate reconstruction of the hologram.

present in the coherent imaging process can later be removed from the digitized wave front data. In particular, if the optical phase error of the lens system can first be measured using a calibrated object source, this error can later be subtracted from subsequent unknown measurements. Unfortunately, BR holograms remain optically sensitive during the readout process. This has the undesirable consequence that the BR hologram is gradually erased by the readout light source. In particular, if not managed diligently, the readout process can contribute more to the hologram degradation than will the effects of thermal decay. As such, it becomes important to minimize this degradation from readout. One method for reducing the effects of readout erasure is to use a longer wavelength. While BR holograms are most sensitive at the blue and green wavelengths, this sensitivity is greatly retarded at the longer, red wavelengths. Consequently, if the BR hologram can be scanned at a longer wavelength, the effects of hologram erasure can be reduced and the amount of recovered information can be increased in proportion (Koek et al 2003). Another method is to efficiently manage the overall exposure of the hologram during the reconstruction process. This can be accomplished in part with a gated reconstruction light source that is synchronized with the CCD camera such that the hologram is only illuminated during the sample-capture cycle of the camera. Methods to reduce the effects of readout erasure will be discussed further in the next section.

4. Displacement measurement using object conjugate reconstruction with BR As an alternative to taking a complete digital scan of the hologram, it is also possible to digitize selected portions of the recorded object space. This section examines such a technique, known as object conjugate reconstruction (OCR) (Barnhart 2001). For some applications that involve displacement measurement, OCR may provide an attractive alternative to the digitization strategies presented previously. In particular, OCR dramatically reduces the required amount of digital information gathered by several orders of magnitude through optical pre-processing of the recorded holographic information. The basic schematic for OCR is shown in figure 6. Initially, as shown in figure 6(a), a double-exposure reflection hologram is recorded of a subject undergoing a displacement by using two identical but laterally displaced converging reference beams at two different time instants, t1 and t2. Then, as shown in figure 6(b), the hologram is reconstructed using a diverging wave from a fibre-optic probe, which is placed in the original object space. The probe, through its three-dimensional position, selects different measurement points of interest within the three-dimensional recorded space. This OCR configuration behaves as an imaging system such that a magnified image of the object space in the region of the probe is produced at two fixed points in space defined by the two previous points of focus of the 643

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Figure 7. Digital holographic displacement measurement from cantilever beam by temporary storage in bacteriorhodopsin and numeric processing via OCR.

recording reference beams (Barnhart et al 2002b). Analogous to methods used in planar PIV (Adrian 1986), the resulting reconstruction introduces a constant shift between exposures that provides a known bias displacement. This image shift is used to resolve any directional ambiguity of the object displacement. As illustrated in figure 6(b), the real image is formed within a pinhole opening, which determines the sample volume of a measurement point. After the optical Fourier transform, the CCD camera detects the two-dimensional spatial power spectrum of the complex field emanating from the aperture (Goodman 1968). Ultimately, each sampled spatial power spectrum contains the necessary information to make a single three-dimensional vector displacement measurement (Coupland and Halliwell 1992). In practice, the digitized spatial power spectrum of each measurement sample is stored as an image file on a computer and its three-dimensional displacement is extracted by numeric processing methods presented elsewhere (Barnhart et al 2002c). To investigate further the ease-of-use of BR films in holography as well as the displacement measurement accuracy of the OCR method, we have chosen to measure the threedimensional displacements along a classic cantilever beam set-up. The out-of-plane displacement results are shown in figure 7. Here, the first four points were taken on the clamp that holds one side of the cantilever beam and the solid line shows the displacement predicted from cantilever theory (Benham and Warnock 1982). Although not shown, OCR also measures the two transverse, in-plane, displacements as well. In this cantilever arrangement, the in-plane displacement components ideally would have a constant value of zero. However, the RMS fluctuations from the zero displacement point enable us to determine the measurement accuracy for the in-plane components as well. The result shown in figure 7 was actually one of 50 independent experimental measurements taken of the same cantilever system. In contrast with the use of wet-processed holographic films, the time savings and ease of use of BR was very impressive. Instead of requiring an hour or more to record, chemically develop and analyse a single experimental result, the complete experimental holographic measurement was obtained in a couple of minutes. In addition, the 644

low grating distortion afforded by BR enabled an improved displacement resolution by a factor of two in comparison with similar OCR experiments previously conducted with conventional silver halide films (Barnhart et al 2002b). In particular, the obtained results had a rms displacement resolution of 0.1 µm in the longitudinal direction (out-ofplane) and 0.05 µm in the transverse directions (in-plane) with a displacement range of ±6 µm. This represents a dynamic range that is greater than 100:1 for the longitudinal component and greater than 200:1 for the transverse components of displacement. Because the principles of OCR can be applied to flow velocity measurement in same fashion as surface displacement measurement (Barnhart et al 2002a, 2004), the present reported results for surface displacement accuracy and dynamic range would also apply to velocity-field measurements using OCR with BR recording. Although the OCR method is well suited for highresolution, three-dimensional displacement measurement, its implementation with BR does have important limitations worth noting. In particular, the readout process for OCR requires that the reconstruction light maintain the same wavelength as the recording laser source. As a consequence, because the BR material remains sensitive to the reconstruction light source, its holographic image will become degraded with continued exposure. Furthermore, the readout process with OCR degrades the hologram image quality more rapidly than the other methods proposed in this paper. It is therefore essential to efficiently manage the overall exposure of the OCR hologram very carefully during the reconstruction process with the use of a synchronized gated reconstruction light source. In addition, the relaxed image sample resolution requirements of OCR (Barnhart et al 2004) can also permit the use of an image-intensifier with the CCD camera to further reduce the required exposure of each image sample. Nevertheless, the inherent light sensitivity of BR will always place a limit on the total number of displacement measurement samples that can be extracted by OCR from a single experiment. While a value for this upper limit has not yet been determined, it appears unlikely that more than a few thousand displacement measurements could be obtained from a particular OCR hologram recording in BR. Although such numbers are certainly less than other holographic analysis methods, this limitation is partially offset by the enhanced ability of OCR to selectively sample specific points of interest within the measurement volume without scanning the entire space. In addition, the enhanced measurement accuracy afforded by OCR is superior to many other holographic techniques since its numeric aperture can be made higher than otherwise possible. Finally, for dynamic experiments that require pulsed laser recording, the OCR method shares the same limits previously discussed for BR in general. In particular, the energy requirements for object illumination are strongly dependent on the holographic application. For example, in the application of holographic velocimetry in fluids and particlefield measurement, to a crude approximation, the object space illumination could require 5–50 mJ cm−2. Therefore, laser power becomes an issue for illuminating a 10 × 10 × 10 cm−3 volume (using a typical size of a silver halide plate), because the total illumination power could be 0.5–5 J. Otherwise, the use of a smaller illuminated object volume may be required.

Bacteriorhodopsin as a high-resolution, high-capacity buffer for digital holographic measurements

5. Applications of holographic measurement with BR This paper considers next three promising applications that could benefit from the holographic measurement with BR. In particular, due to its combined potential for ease of use and high spatial resolving power, BR holographic measurement could be well suited for the areas of biomedicine, displacement/velocity mapping in fluids and on surfaces, and supersonic measurement. The first to be considered is the application of such an instrument to biomedical applications. In the area of flow measurement and velocityfield mapping, digital holographic measurement could have applications to the study of blood flow and the development of vascular prosthetics including artificial heart valves. A related area of research is in the flow measurement of the air movement within the lungs. In the area of microfluidics technology, this instrument can be used to obtain high-resolution, quantitative, velocity-field measurements of the microscopic, three-dimensional flows within such devices. In some cases, a holographic camera based on BR could be fitted to capture multiple hologram frames of information at several different instants in time using a method of angular/spatial reference-beam multiplexing. This would enable the holographic recording of the evolution of a process over time and the measurement of velocity fields in space (Hinsch 2002). Once all such hologram frames have been recorded, each frame can then be sequentially scanned and digitized by the methods proposed in section 3 of this paper. Another promising application is in its use as a holographic microscope for high-resolution, high-volume imaging of cell-to-cell interactions, cell population dynamics and cell membrane adhesion. In particular, a digital holographic system could be used to record, across centimetrescale volumes with sub-micron resolution, the simultaneous behaviour of millions of living cells. If the information is digitally recorded and numerically reconstructed, it also might be possible to use artificial-vision computer algorithms that recognize the locations and types of different cells and cell behaviours in order to statistically determine the cell population dynamics. A related topic of study is the cellto-cell interactions of developing embryos. Because the digital holographic microscope records information over a large volume, it becomes possible to observe living cellular organisms as they are functioning in their natural environment without any artificial physical constraints caused by the cell containment or chemical stains that could inhibit the natural cell behaviour. Because the holographic microscope records the optical phase information, transparent objects could be viewed without difficulty. One final application worth mentioning is the holographic measurement of supersonic events. When velocities exceed 1000 m s−1, holographic measurements require ultrashort exposures in order to freeze the recorded interference fringes. In such instances, exposure durations of less than 100 ps may be required. Unlike most other recording materials such as photographic film and electronic cameras, BR film actually exhibits improved light sensitivity under such circumstances (Hampp et al 1990).

6. Conclusion This paper has reported on the use of bacteriorhodopsin (BR) for holographic recording with both optical and numeric reconstruction. One of the greatest benefits of BR-based holography is the fast turnaround time for an experimental measurement. Rather than requiring hours to conduct an experiment with wet-processed holography, the same experiment with BR holography can be accomplished in minutes. In addition, in many respects, bacteriorhodopsin (BR) appears to be well suited for digital holography. In contrast with direct digital holography on CCD cameras, BR holograms are not restricted to microscopic object dimensions but, rather, can take volumetric measurements comparable with traditional silver halide holograms. Nevertheless, the BR material does possess limitations that must be carefully managed. Unlike the inherent tolerance of silver halide holography to varying degrees of light exposure, BR holography demands careful controlling of the reference beam energy. Another limitation of BR holograms is the limited life of the recorded fringe information (but not the material itself). This limited time span may also restrict the amount of information that can be recovered from the hologram. Finally, for dynamic object measurements, the available energy capacity of the pulsed laser can place an upper limit on both the size of the hologram aperture as well as the object space volume. Nevertheless, once these limitations have been taken into consideration, it appears that the use of BR could be highly effective, in terms of both quality and process time, for a wide range of three-dimensional measurements. Consequently, several holographic applications, based on BR, have been proposed in this paper.

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