In the USA and other developed countries, cancer is responsible for 25% of all deaths. In spite of the most recent advances in cancer research, by far the best curative treatment for the disease is early detection. Such technology revolves around quick identification of malignant cells, necessitating clean, crisp images for oncologists. The CellTracks Analyzer II by Immunicon (Huntington Valley, PA) is a semi-automated fluorescence microscope system used to count and characterize fluorescently labeled cells that are immuno-magnetically selected and aligned. The SEBS-B type miniature slide guide, from NB Corp. of America (Wood Dale, IL), is tapped as part of the microscope’s system of axes.
Cancer is a growth of cells that is unrestricted and inchoate. As a term, “cancer” usually refers to malignant neoplasm only, those cancers characterized by spreading to other sites in a process called metastasis. In a metastatic invasion, cancer cells break off of the primary tumor and travel to remote locations in the body via the blood stream, lymphatic system, or both and there take root in the form of a new tumor. While metastasis is a characteristic of late-stage tumor growth, doctors and patients usually do not take the chance at any stage of the disease. Detection methodologies, from mammography to the CellTracks Analyzer, must be able to clearly differentiate cancerous cells from a background of otherwise healthy cells that number in the billions. The CellTracks Analyzer II tests the blood of the patient, searching for circulating tumor cells (CTC) that have since broken away from the primary tumor and flowing through patient’s bloodstream.
A fluorescence microscope is a light microscope used to study properties of substances using fluorescence and/or phosphorescence rather than, or in addition to, the reflection and absorption techniques of conventional microscopes. The light used is also usually more intense than convention counterparts.
The CellTracks Analyzer II, designed to count and characterize CTCs, contains several NB components; the stainless SEBS-B guides are found in three (of the four) axes, consisting of a block and guide rail, both of which have two precision ground raceway grooves. Wanting to keep the design compact, Immunicon had to pay close attention to the size of all the components used. The two-raceway and four-point contact structure of the SEB-B minimizes their height, making them suited for the limited space provided; the SEBS is the smallest and lightest slide guide series in NB’s stock. Due to the need for fine precision for the viewer, the ability of the guides to maintain a constant level of friction and running (Immunicon used the nonretained-ball version of the SEBS-B) was a determining factor in the CellTracks’ design. Immunicon choose the SEBS-B specifically for its preload; for the fact that it has no “play.” Two axes using the slide guide control the movement of the sample, requiring an unvarying smoothness of motion. The third SEB-S axis moves one of three optical blocks, called filter cubes, into position in the optical path of the microscope. This cube provides a convenient means to change the microscope’s mirror without direct handling of either the mirror or filters.
The CellTracks Analyzer II is a type of epi-fluorescence microscope, where observation of fluorescence are from above (“epi”) the sample. Like a regular microscope, a fluorescent microscope produces a magnified image of the sample, only the image emanates from fluorescence of the subject, rather than from the light originally used to illuminate the sample. As a unit, CellTracks Analyzer II consists of a dedicated computer loaded with CellTracks software, mouse, and keyboard. Immunicon uses a “systems” approach to cancer detection. The CellTracks included a tube developed to preserve blood, presumably with CTC cells, en route to a lab. The device contains a system that standardizes and automates sample preparation. Images of all filters are compiled and presented in a gallery format for final cell classification by a lab technician on the computer. In a typical epi-fluorescence microscope, the illumination light (with regard to the CellTrack Analyzer II, from a mercury arc-discharge lamp) first filtered of unnecessary wavelengths and bounced off a dichroic mirror (a type of mirror that reflects some wavelengths but allows others to pass through it) to the sample. Light coming from the sample passes through the dichroic mirror again, and is then separated further from the much weaker emitted fluorescence via an emission filter. The dichroic mirror is mounted to the filter cube. The excitation and emission filters are usually also affixed to the filter cube.
In a typical protocol is as follows: a blood sample is collected in the tube and placed in a cartridge containing a reaction chamber. The cartridge, with the sample, is inserted into a cell presentation device, a fixture of two magnets held together by steel. Cells are then subjected to magnetic nanoparticles, called ferrofluids, that are conjugated to antibodies that target rare cells including CTCs. As part of the cell isolation process, a separate device that is part of the CellTracks system, called a MagNest, magnetizes the sample. This causes the targeted cells to become immuno-magnetically labeled — targeted cells that have the ferrofluids bound to them via the antibodies collect in the magnetic field. The magnetized cells are also brought closer to the surface of the reaction chamber, ready to be scanned. The magnets are then retracted, the cells are re-suspended, and staining reagents are added. Staining (also called labeling), involves target-specific, fluorescent dyes called fluorophores. In the case of the CellTracks Analyzer, three fluorophores are used, one to color the nucleus of the cell, another stains the cell membrane and other non-nucleic structures, and another colors specifically white blood cells. Cancerous cells are identified when they register with the first two dyes, but not the third. The specimen is then illuminated with light at a specific wavelength that is absorbed by the fluorophores, causing them to emit longer wavelengths of light; a common fluorophore, GFP, is harvested from jellyfish, and fluoresces a characteristic bright green when exposed to blue light. The process is very specific, allowing doctors to find a relatively small amount of targeted cells, or even one, among several billions of others.