Biomedical Engineering Reference
In-Depth Information
Secondary oocyte
Cone of rays
that is traced
Model of the
mouse oocyte cell
Δ
n
1
Δ
n
2
Zona pellucida
Scan
40
μ
m
FIGuRE 6.4
Transmitted.light.image.of.the.mouse.oocyte.cell.(let).and.the.geometric.model.used.for.the.simula-
tion.of.wavefront.aberrations.(right).
real.cell.had.a.diameter.of.approximately.80.μm,.and.a.range.of.100.μm.was.scanned.on.a.16.×.16.grid..he.
simulation.calculates.the.ield-dependent.component.of.the.aberration.only..Additional.aberration.terms.
due.to.the.refractive.index.mismatch.between.immersion.medium.and.embedding.medium.are.not.consid-
ered.here.but.have.been.calculated.by.Török.et.al..(1995).and.Booth.et al..(1998)..he.simulation.assumed.
diameters.of.the.two.concentric.spheres.of.80.and.52.μm.as.inferred.from.a.transmitted.light.image.of.the.
cell.and.used.the.values.for.the.NA.(dry.lens,.NA.=.0.5).and.the.“scanned”.ield.(100.μm.×.100.μm).as.deter-
mined.from.the.experiment..he.diference.in.the.values.of.refractive.index.between.embedding.medium.
and.the.biological.specimen.could.not.be.determined.by.a.complementary.method,.but. ∆ =
n
1
0 016
.
.and.
n
2
0 03.
.(inner.sphere).were.found.to.indicate.a.good.agreement.with.the.experimental.data..In.the.
experiment,.the.focus.was.set.to.the.supposed.bottom.of.the.cell.with.an.accuracy.better.than.5.μm..A.set-
ting.of.10.μm.above.the.lower.edge.of.the.outer.sphere.of.the.simulation.was.found.to.give.the.best.reproduc-
tion.of.the.experimental.data..his.could.be.explained.by.the.fact.that.the.cell.is.not.perfectly.spherical.and.
the.cross.section.in.the.
z
-direction.is.smaller.than.that.in.the.
x
-direction.
Figure.6.5
.shows.the.simulation.and.the.interferometric.wavefront.measurement.results.next.to.each.
other.. he. Zernike. mode. charts. for. both. the. data. sets. are. displayed. at. the. same. scale. and. show. the.
variation.of.the.diferent.modes.across.the.scanned.ield.of.view..he.experimental.setup.uses.a.phase-
stepping.interferometer.in.transmission.geometry.in.which.the.sample.is.mounted.between.two.oppos-
ing.objective.lenses.contained.in.the.measurement.arm.of.the.interferometer..A.mirror.mounted.in.the.
reference. path. is. moved. by. a. piezo. actuator. and. performs. phase. stepping. at. videorate.. he. sample. is.
mounted.on.an.
x-y
.stage.and.translated.perpendicular.to.the.optical.axis.and.wavefronts.are.recorded.
on.a.16.×.16.grid.across.the.specimen..For.experiment.and.simulation,.Zernike.modes.2.through.22 were.
extracted. from. the. wavefronts. but. the. igures. show. modes. 2. through. 12. only. since. the. higher-order.
modes.have.relatively.low.amplitudes..It.should.be.mentioned.that.an.adaptive.optics.aberration.cor-
rection.system.would.not.correct.the.Zernike.modes.tip.(2),.tilt.(3),.or.defocus.(4).since.these.aberration.
modes.correspond.to.lateral.or.longitudinal.translation.of.the.focal.spot.but.leave.signal.intensity.and.
resolution.unafected..he.presence.of.these.modes.in.the.aberrated.wavefront.causes.geometric.distor-
tions.in.the.obtained.3D.data.set.because.there.is.a.diference.between.the.actual.and.the.predicted.posi-
tions.of.the.focal.spot..However,.correction.would.require.a.detailed.knowledge.of.the.refractive.index.
distribution.of.the.specimen.
Note.that.the.experimental.measurement.shown.in.
Figure.6.5
.
contains.the.static.aberration.compo-
nent.caused.by.the.refractive.index.mismatch.in.addition.while.the.simulation.covers.the.ield-dependent.
fraction. only.. he. static. component. mainly. afects. mode. 11. (irst. spherical). and. causes. an. additional.
Zernike.amplitude.ofset.for.this.mode..he.agreement.between.experiment.and.simulation.is.good,.and.
the.main.features.of.the.Zernike.mode.variations.across.the.ield.are.reproduced.by.the.simulations.
∆ =
