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Phase Contrast Test Slide

H.S.E Test Slide Image, 40X NA 0.65 Phase Contrast Objective

This is the pattern of lines on the standard Phase Constrast Test Slide used to assess the sensitivity to phase objects when using phase constrast microscopy. This photograph shows groups 4 through 6 using a 40X phase contrast objective, NA 0.65,using a microscope (Olympus BH2) used by many contractors assessing the presence of airborne asbestos fibers.

Transmitted Phase Contrast Illumination, 40X Objective, NA 0.65


The Phase Contrast Test Slide is manufactured to test the sensitivity of a phase contrast microscope to a given shift in phase between the direct ray and the diffracted ray paths for a specific configuration of the microscope. This slide is designed to test sensitivity to phase and not the resolution limit or limit of visibility for the microscope. This photograph shows that the groups of groves #4 through #6 are visible using this light intensity (setting of 6 volts). Neutral density filters can be added to decrease the constrast sufficiently to meet the specification requirments or the microscopist can ignore the #6 group of lines and learn to get the same counts that agree with the majority of the other laboratories.

Significance in the Environment:

This slide has been represented incorrectly by some as a test of the resolution limit of the microscope in a given configuration. It is actually used as a way to calibrate the microscopist to some vague average value for given round-robin test samples. The ability to see asbestos fibers on a cleared filter are a combination of the quality of the optics (microscope) and the capability of the microscopist. The range of visual acuity varies significantly from individual to individual and for a given individual with current fatigue level and age. Add to this the difference in experience and the microscopist can be seen as a major variable in the results.

Characteristic Features:

The patern on this slide consists of seven bands of twenty lines each. The width of the individual lines range from 1.1 micrometer for the first set to 0.25 micrometers for the seventh set of lines. The depth of each engraved line from which the resin cast is made is approximately one tenth of the line width. The refractive index of the resin used to make the cast is 1.58.

Associated Particles:


LeGuen, J.M.M., T.L. Ogden, T. Shenton-Taylor and J.F. Verrill, The HSE/NPL phase-contrast test slide", Ann. Occup. Hyg., Vol. 28, no. 2, pp. 237-247, 1984 This reference briefly gives some of the specifics on the design of the test slide.