Use of Field Finders

Locating a small area of special interest on a glass slide (e.g., a particular clean region of well-spread individual sperm nuclei), can be a tedious task of scanning a large area of the slide to reposition that same field on the microscope for cytophotometry. Ink marks or circles marked with a wax pencil are sometimes difficult to apply and often rub off. The job can be made much simpler by the use of a slide marked by a numbered and lettered grid (see Fig. 15). With this tool, it is possible to locate a particular group of cells and record their coordinates in terms of space designations on the surface of a 3 x 1-in. glass slide called a "field finder" (see Note 18). Gridded slide finders are marked so that they are read right side up (to the viewer) in a conventional microscope when inserted into the clasping jig of the mechanical stage. It is important to position both the "test" slide (the slide with the cells that you want to relocate to measure them) and the gridded slide very carefully so that the corners of the slides are fitted securely within the flat holding arms of the mechanical stage so that repositioning can be accomplished readily and repeatedly without displacement because of stage backlash. To do this, perform the following:

1. Fit the test slide into the clasp of the mechanical stage, being certain that the slide fits snugly against both the vertical and the horizontal axes and the crescent-shaped holding arm of the stage.

2. Scan the slide to find the objects of special interest.

3. Taking care not to disturb the position of the mechanical stage, remove the "test" slide and replace it with the gridded slide. Focus on the field finder without moving or touching the mechanical stage. Note both the vertical and horizontal coordinates of the grid location, reading the site for the position of the cells that you positioned previously as both a letter and a numerical value, noted as a decimal of the letter (e.g., S.2), as shown in Fig. 15.

Fig. 15. The Lovins Field Finder (A) and the England Finder Grid (B) are precision devices for the microscopist to use in relocating fields of interest on a slide-mounted specimen. Each finder has a precision rectangular-coordinate grid pattern protected by a thin cover glass on a special microscope slide. (See text and Note 18 for more details.) The appropriate answers for grid locations of symbols with the Lovins Finder are given at the upper left of Fig. 14. The location of the asterisk in the lower left quadrant of the England Finder field of view is read as 34/3*. It does take practice and a bit of time to learn how to read slide coordinates for your prized slide preparation with either of the finders, but the time spent to master this game is a small investment when compared to the time spent trying to relocate a particular group of cells that you know is on that slide.

Fig. 15. The Lovins Field Finder (A) and the England Finder Grid (B) are precision devices for the microscopist to use in relocating fields of interest on a slide-mounted specimen. Each finder has a precision rectangular-coordinate grid pattern protected by a thin cover glass on a special microscope slide. (See text and Note 18 for more details.) The appropriate answers for grid locations of symbols with the Lovins Finder are given at the upper left of Fig. 14. The location of the asterisk in the lower left quadrant of the England Finder field of view is read as 34/3*. It does take practice and a bit of time to learn how to read slide coordinates for your prized slide preparation with either of the finders, but the time spent to master this game is a small investment when compared to the time spent trying to relocate a particular group of cells that you know is on that slide.

4. Put the slide finder into position on the mechanical stage of the microscope to be used for cytophotometry and relocate the letter-number locus.

5. Remove the field finder and position the "test" slide into the mechanical stage and focus for the object plane without touching or moving the mechanical stage.

6. Again, focus under low power for the object plane, without moving the mechanical stage to identify the group of nuclei to be measured. They should now appear within a 20- to 25-|jm radius of the center of the field and might require slight repositioning to go to a 40x high dry objective or to a 100x oil immersion lens for measuring and/or photography.

7. Locate other sites of interest for measuring on the same or other slides by repeating steps 1-5 with the "test" slide and then the field finder.

Often, it is sufficient to determine the object locus to a letter and one decimal point before putting the test slide on the stage of the cytophotometer using the coordinates of the gridded slide to position the "test" slide at the same position as the gridded slide that had been used to relocate the desired field. It is also possible to use the letter locus with an estimate of the second decimal point for relocation of a site on the "test" slide initially under low power, before going to oil immersion for final location of the object of interest. This also makes it feasible to find suitable areas for background readings near the nuclei to be measured. To clean a gridded slide, wipe it with a soft cloth or lens paper. Avoid solvents or other chemical cleaners.

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