Tech Briefs

With careful planning, system integrators can select the optimal optics, filters, light sources, and cameras for their medical diagnostic instrumentation.

Custom integration of original equipment manufacturer (OEM) products can be complex, particularly for medical device integrators that build diagnostic instruments incorporating numerous optical components. Often, objective lenses, illumination sources, and imaging detectors are assembled and custom-mounted into finished instruments. Such components must not only meet stringent performance requirements, but often have to meet established Food and Drug Administration (FDA) standards.

Fig. 1 – Human Breast Cancer tissue, stained in FISH (fluorescence in situ hybridization) technique (used to detect and localize the presence or absence of specific DNA sequences on chromosomes).

Medical device design and manufacture includes traditional areas, such as the manufacture of stents, surgical implants, and other products used to diagnose and treat patients, along with instruments that aid scientists in areas such as genetic sequencing and cancer detection. Some devices use transmitted light microscopy and are able to employ advanced technology such as virtual microscopy or advanced cellular imaging techniques to provide images used by pathologists for diagnosis.

As performance, reliability, and other requirements evolve, the imaging field is changing and technology is advancing rapidly. In many cases, instruments have become highly controlled environments featuring self-contained, turnkey imaging systems that require minimal operator control. Automated optical components may be used to find focus quickly and repeatedly, making it possible for manufacturers to set up systems that are programmed to record image data for every sample.

Light Considerations: Performance, Objectives

Fig. 2 – Numerical aperture is one of the key objective lens specifications, referring to the light gathering ability of the lens and depends on the refractive index (n) of the immersion media.

Performance specifications such as resolution, magnification, and transmission are vitally important for systems with optical components or microscopes. Lateral resolution refers to the smallest distance between two points on a specimen that can be distinguished as two separate entities — for example, distinguishing between two cells or intracellular features. Numerical aperture (NA), one of the most important specifications for any objective lens, refers to the light-gathering ability of the lens. Resolving power increases proportionally with NA and, generally speaking, higher-NA objectives carry a higher price tag as well.

Most objective lenses are designed for peak performance at a specific wavelength defined by transmission characteristics and chromatic aberration. Match ing system wavelengths to the appropriate objective is critical to overall performance. Many objective lenses are corrected for the visible light spectrum (400–700 nm). Other specialized objectives peak in the ultraviolet (UV700nm) ranges.

Fluorescence is one of the most useful techniques for imaging subcellular structures. Particular portions of a cell are tagged with a fluorescent label, which causes them to emit light of a specific longer wavelength when stimulated by photons of a particular shorter wavelength. When specifying systems for fluorescence and related imaging methodologies, additional components may be required, including filter cubes that specifically match the illumination source and objective. These cubes are paired with excitation and emission filters specifically designed to match the fluorescent tag.

One method of improving resolution in cell biology, anatomic pathology, cytopathology, and other fields is the use of immersion objective lenses. Resolution is dependent on numerical aperture defined by the equation NA = n (sin μ), where n is the refractive index of the medium between the objective and the specimen. The maximum NA using a dry objective is 0.95, where n=1. By using immersion media with a refractive index greater than 1, the NA of the optical system can be increased. As an example, typical immersion oil has a refractive index of 1.518, which will increase the NA to as high as 1.4. To use an immersion objective, the microscopist lowers the tip of the objective into water, specialized oil, silicone or glycerine, depending on the refractive index desired, and images the sample directly through the immersion medium. Immersion optics often requires the use of a coverslip and a specially designed correction collar that is used to adjust for the thickness of the sample.

Mechanical and Illumination Considerations

In specifying optics, mechanical and illumination considerations are also crucial. Systems may need to deliver a specified spatial resolution for optimal visual contrast when imaging or collecting spectral data. Most optical systems today use an infinity-corrected optical design, which allows intermediate modules and components to be introduced without affecting magnification. There is, however, a fixed range for the distance between the objective lens and tube lens.

Working distance (WD) is related to the utility and design of optical systems. It refers to the distance between the glass element of an objective’s mounting structure and a focused specimen. The longer the WD, the more space there is between the objective and the focused image plane. This provides two benefits — sample safety (the objective must travel further before it contacts the sample) and the ability to image a sample located within a defined package or holder such as a Petri dish or well plate. Typically, however, an increase in WD corresponds to a decrease in NA.

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