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The onset of nanotechnology and targeted therapy methods for a number of pathologies has made it increasingly more difficult to image effectively in the medical field. With that being said, the inception of quantum dots and the improvements to optical filters has made this once daunting task a common practice.
Stephan Briggs, Edmund Optics
The onset of nanotechnology and targeted therapy methods for a number of pathologies has made it increasingly more difficult to image effectively in the medical field. With that being said, the inception of quantum dots and the improvements to optical filters has made this once daunting task a common practice. The appealing nature of quantum dots in conjunction with optical filters is the ability of quantum dots to be tagged to specific biological agents for long durations of time and then imaged via the fluorescent energy they emit. This fluorescent energy that is emitted is selectively filtered resulting in a crisp and pristine image that can easily differentiate between a normal, healthy cell and a deadly, malignant cell. One prime example is the imaging of Reed-Sternberg cells that are common in cases of Hodgkin's lymphoma. Typically an easy cell to identify, but in the case of Hodgkin's lymphoma are surrounded by a plethora of healthy white blood cells and a deep bed of other lively tissues (1).
Through spectroscopy one can effectively quantify the interaction between light energy and matter. There are many types of spectroscopy which include absorption, X-ray, visible, ultraviolet, infrared, fluorescence, and many more. As mentioned above, fluorescent spectroscopy can be utilized in medical imaging systems efficiently as photons excite the sample of interest. In this case, the sample of interest is a quantum dot actively targeted for a cancerous region which then emits a low level of energy that can be detected with a spectrometer. In the detection of Reed-Sternberg cells, it is crucial to understand which wavelength is being emitted at which region. One must differentiate between a metastatic region and a healthy tissue bed without error otherwise there will be unnecessary cell death and tissue damage. To do this best and most effectively spectroscopy is put into action.
Fluorescent microscopy systems are becoming more and more of an industry standard in the medical field. These systems typically consist of a few entities — excitation, emission and dichroic filter, illumination, and an imaging sensor. The dichroic and emission filter are the two most crucial elements. Together they prevent non-emission energy and stray light from reaching the sensor. In this type of microscope system, the quantum dot in place absorbs the excitation energy and then converts it to a radiating glow, or rather emission energy.
The important parameters of a filter include the center wavelength (CWL), minimum transmission percentage, optical density (OD), and bandwidth, which at times can also be referred to as the full width at half maximum (FWHM). The CWL and bandwidth work together and refer to the wavelength where maximum transmission is reached and the region to which 50% transmission is still attainable. The minimum transmission dictates the amount of attainable fluorescence and excitation from the quantum dots. This value should be as high as possible, with an industry standard at or around 85%. New and improved filters that have increased blocking efficiencies are now achieving >93% minimum transmission. The OD refers to the filter's blocking efficiency to light and wavelengths outside the desired bandwidth. With a higher OD there is improved reduction of noise and a sharper cutoff between transmission and rejection, resulting in sharper colors and higher contrast. Lastly, the final element of importance is the dichroic filter. These filters are specifically in place for cleanup of stray and excess light. Ideally, they will possess an extremely crisp transition between maximum reflection and maximum transmission. The point of maximum transmission will ensure minimal stray-light and a maximum signal-to-noise ratio which is crucial for fluorescence imaging.
The spectral characteristics of the quantum dot in place dictate which filters would work best in the fluorescent microscope system. Compared to a typical fluorescent agent, quantum dots are proven to be much brighter and more stable, by a scale of 20× and 100× (3). Not only will quantum dots fluoresce and emit a much higher energy, but there is very little risk of photo-bleaching by the illumination source as they are synthesized from semiconductor materials. The two key parameters are the peak excitation λ (nm) which refers to the wavelength the quantum dot absorbs best, and the maximum wavelength that it then emits after absorption, which is known as the peak emission λ (nm). A third parameter is the excitation range of the quantum dot, which typically resides within the FWHM of the filters. Larger quantum dots have closely spaced energy levels, and therefore would emit a small band of wavelength, whereas smaller dots have more widely spaced energy levels, and would ultimately emit a much larger wavelength band (2).
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(1) C.T. Troy, "Multicolor quantum dots ID rare cancer cells." bioPhotonics 10 (2010).
(2) A.F. Van Driel, "Frequency-dependent spontaneous emission rate from CdSe and CdTe nanocrystals: Influence of dark states." Physical Review Letters 95 (2005).
(3) M.A. Walling and S. Novak, "Quantum dots for live cell and in vivo imaging." International Journal of Molecular Science 441 (2009).
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