Problems in polarized light microscopy observation of birefringence of calcium pyrophosphate dihydrate crystals

To reconsider the problems arising from the use of the phase plate as a test plate inserted into a polarized light microscope system for the analysis of triclinic calcium pyrophosphate dihydrate (t-CPPD) crystals, or Ca2P2O7·2H2O in the synovial fluid of arthritis patients, we made the polarized light microscopy observations using a phase plate with a retardation of 530 nm for the synthesized t-CPPD crystals well-characterized by X-ray powder pattern indexing and single crystal X-ray diffraction measurements. The microscopy observations were made of crystals of different sizes, thicknesses and shapes. The retardation was assessed using the interference color chart at four extinction and diagonal positions both with and without the test plate. The addition and subtraction states produced by superimposing the retardations of two anisotropic materials, that is, the t-CPPD crystal and the 530 nm phase plate, were deduced from the interference color change by inserting the test plate at four diagonal positions. When the color change of a crystal at a diagonal position resulting from 90-degree rotation exhibits no clear birefringence, the interference color chart was shown to be useless. We suggested the use of a compensator whose retardation can be changed to obtain an accurate value for the retardation of the crystal.