Aberrations and the Benefit of Their Correction in Confocal Microscopy - Adaptive Optics for Biological Imaging - page 64

inset of Figure 4.13 . Note that again the variation of the coeicients declines with increasing order of
the successive Zernike modes. As with the previous sample, coeicient 11 (spherical) shows a signiicant
ofset, as expected. For both the mouse specimens, the objective lens Zeiss Plan-Neoluar 20×, 0.5 NA
was used. his dry lens does not have a coverslip correction, but the correction ring of the condenser
lens was adjusted during the calibration. herefore, the additional spherical aberration introduced by
the coverslip of the sample was compensated. Only the spherical aberration originating from the liquid
layer of the specimen was added to the total aberration measured.
For the last sample, a C. elegans specimen of cylindrical shape, the experimental conditions were
slightly diferent: an Olympus 20×, UPlanApo, 0.8 NA, oil immersion was used as the objective lens, and
a one-step calibration in respect to the reference wavefront 2 within the specimen slide was carried out.
his means that the aberration data shown in Figure 4.12 and the bottom inset of Figure 4.13 contain
the ield-dependent fraction of the specimen-induced aberration only; the static part caused by focusing

Total Zernike Mode 2—Tip Mode 3—Tilt

1 1

0.75 0.5

0.75 0

0.25 ‒ 0.5

0 ‒ 1

Mode 5—Astigmatism Mode 6—Astigmatism Mode 7—Coma

Mode 4—Defocus

1

0.5

0

‒ 0.5

‒ 0

Mode 8—Coma Mode 9 Mode 10 Mode 11—Spherical (1st) Mode 12

Mode 13 Mode 14 Mode 15 Mode 16 Mode 17

Mode 18 Mode 19 Mode 20 Mode 21 Mode 22—Spherical (2nd)

Figure 4.12 Zernike mode plots of the C. elegans sample, coeicients 2-22. For these images, the Zernike modal
content was extracted from 256 wavefronts. Objective lens: Olympus 20×, UPlanApo, 0.8 NA, oil immersion.
Condenser lens: Zeiss LD-Achroplan, dry lens, 40×, 0.6 NA, correction ring. he scanned area was 100 × 100 μm.