Colin J. R. Sheppard

Electro-spinning of pure collagen nano-fibres - just an expensive way to make gelatin?

Scaffolds manufactured from biological materials promise better clinical functionality, providing that characteristic features are preserved. Collagen, a prominent biopolymer, is used extensively for tissue engineering applications, because its signature biological and physico-chemical properties are retained in in vitro preparations. We show here for the first time that the very properties that have established collagen as the leading natural biomaterial are lost when it is electro-spun into nano-fibres out of fluoroalcohols such as 1,1,1,3,3,3-hexafluoro-2-propanol or 2,2,2-trifluoroethanol. We further identify the use of fluoroalcohols as the major culprit in the process. The resultant nano-scaffolds lack the unique ultra-structural axial periodicity that confirms quarter-staggered supramolecular assemblies and the capacity to generate second harmonic signals, representing the typical crystalline triple-helical structure. They were also characterised by low denaturation temperatures, similar to those obtained from gelatin preparations (p>0.05). Likewise, circular dichroism spectra revealed extensive denaturation of the electro-spun collagen. Using pepsin digestion in combination with quantitative SDS-PAGE, we corroborate great losses of up to 99% of triple-helical collagen. In conclusion, electro-spinning of collagen out of fluoroalcohols effectively denatures this biopolymer, and thus appears to defeat its purpose, namely to create biomimetic scaffolds emulating the collagen structure and function of the extracellular matrix.

Image Formation in the Scanning Microscope

Fourier imaging in the scanning microscope is considered. It is shown that there are two geometries of the microscope, which have been designated Type 1 and Type 2. Those of Type 1 exhibit identical imaging to the conventional microscope, whereas those of Type 2 (confocal microscopes) display various differences. Imaging of a single point object, two-point resolution and response to a straight edge are also considered. The effect of various arrangements using lenses with annular pupil functions is also discussed. It is found that Type 2 microscopes have improved imaging properties over conventional microscopes and that these may be further improved by use of one or two lenses with annular pupils.

Advances in optical and electron microscopy

J-P Adriaanse High-Te Superconductors and Magnetic Electron Lenses. E. Kohen and J.G. Hirschberg, Microspectrofluoremetry. S. Kawata, Optical Computed-Tomography Microscope. Y. Shimizu and H. Takenaka, Microscope Objective Design. Subject Index.

Annular pupils, radial polarization, and superresolution

An annular pupil, which can be used to produce a Bessel beam, when combined with radially polarized illumination promises improvements in microscope resolution, increased packing density for optical storage, and finer optical lithography. When combined with a circular detection pupil in confocal microscopy a point-spread function 112 nm wide results (lambda = 488 nm). Radially polarized annular illumination of a solid-immersion lens can yield a focal spot smaller than 100 nm for lambda = 488 nm. Use of radially polarized illumination with pupil masks is discussed.

Optical image encryption based on diffractive imaging

In this Letter, we propose a method for optical image encryption based on diffractive imaging. An optical multiple random phase mask encoding system is applied, and one of the phase-only masks is selected and laterally translated along a preset direction during the encryption process. For image decryption, a phase retrieval algorithm is proposed to extract a high-quality plaintext. The feasibility and effectiveness of the proposed method are demonstrated by numerical results. The proposed method can provide a new strategy instead of conventional interference methods, and it may open up a new research perspective for optical image encryption.

Linear phase imaging using differential interference contrast microscopy.

We propose an extension to Nomarski differential interference contrast microscopy that enables isotropic linear phase imaging. The method combines phase shifting, two directions of shear and Fourier‐space integration using a modified spiral phase transform. We simulated the method using a phantom object with spatially varying amplitude and phase. Simulated results show good agreement between the final phase image and the object phase, and demonstrate resistance to imaging noise.

Depth of field in the scanning microscope.

Various definitions of depth of field in the microscope are discussed. The variation in the integrated intensity in the image of a point object outside the focal plane shows how the microscope discriminates against such objects. The power diffusely scattered by a translucent object is also considered. A Type-2 scanning microscope is found to have a much reduced depth of field according to these criteria, which makes it useful for studying thick biological slices. These results do not contradict the claim that depth of field may be much increased in such a microscope by using lenses with annular pupil functions.

Information capacity and resolution in an optical system

The concept of invariance of information capacity is discussed and applied to the resolution of an optical system. Methods of obtaining superresolution in microscopy are discussed, and scanning microscopy has many distinct advantages for such applications.

Imaging in high-aperture optical systems

Nonparaxial effects on optical imaging are considered. It is shown that the dominant effect is on the axial variation in the complex amplitude, affecting most strongly depth of focus, interference microscopy, and the imaging of objects with appreciable depth. A so-called pseudoparaxial approximation is introduced, which gives reasonable prediction of these effects up to quite large angular apertures.

Second-harmonic imaging in the scanning optical microscope

A scanning optical microscope in which an image is produced from the generation of optical second harmonics within the specimen has been constructed. Pictures have been obtained from various crystals which show high contrast levels and detail not visible with the conventional microscope.