DEVELOPMENT OF A NARROW WATER-IMMERSION OBJECTIVE FOR LASERINTERFEROMETRIC AND ELECTROPHYSIOLOGICAL APPLICATIONS IN CELL BIOLOGY

Laserinterferometric studies of the micromechanical properties of the organ of Corti using isolated temporal bone preparations are well esta blished. However, there are relatively few measurements under in vivo conditions in the apical region of the cochlea because of its inaccess ibility with commonly used techniques. Recently, optical-design progra ms have become affordable and powerful, so that the development of an optimized optical system is within the budget of physiologists and bio physicists. We describe here the development of a long-range water-imm ersion objective. To circumvent anatomical constraints, it has a narro w conical tip of taper 22 degrees and diameter 2.4 mm. It is a bright- field reflected-light illumination, achromatic objective with magnific ation of 25 x/infinity, a working distance of 2.180 mm and a numerical aperture of 0.45. Chromatic errors are corrected at 546.1 and 632.8 n m, with emphasis on the latter wavelength which is used by the laser i nterferometer. The field curvature is relatively flat and a diffractio n limitation (Strehl ratio better than 0.8) can be obtained in a field of 0.4 mm diameter. Using this objective, sound-induced vibrations of hair cells and Hensen cells could be recorded without placing a refle ctor on the target area. In addition, this objective was found to be d iffraction-limited in the near infra-red (750-830 nm), with a slightly different working distance (2.186 mm), making it suitable for patch-c lamp experiments using infra-red, differential interference contrast. (C) 1997 Elsevier Science B.V.