Adaptive Optics in Wide-Field Microscopy - Adaptive Optics for Biological Imaging - page 292

46 seconds.on.a.Core.2.Duo.Intel.processor..Figure.15.23.shows.the.improvement.in.peak.intensity.and.
Strehl.ratio.for.the.data.of. Figure.15.22 ..Figure.15.24.compares.the.width.of.the.corrected.PSF.to.the.
ideal.image.of.a.200.nm.bead..he.measured.PSF.has.an.FWHM.of.290.nm,.about.20%.greater.than.the.
ideal.case..We.performed.the.same.measurement.but.taking.only.ive.slices.per.image.stack.with.a.step.
of.1.0.microns..Taking.fewer.steps.speeds.up.the.phase.retrieval.and.also.reduces.the.light.exposure.of.
the.sample,.but.results.in.less.robust.phase.retrieval..In.this.case,.the.Strehl.ratio.increased.from.0.004.
to.0.30,.and.the.intensity.increased.by.a.factor.of.8.2.in.four.cycles.

15.3.5.1 Phase retrieval from Biological Samples
Phase.retrieval.can.also.work.on.point.sources.embedded.in.or.below.biological.samples.. Figure.15.25 .
shows.phase.retrieval.from.a.200.nm.luorescent.bead.embedded.in.a. Drosophila .embryo..he.dominant.
Zernike. mode. of. the. retrieved. phase. is. astigmatism. (1.5. radian. RMS. amplitude),. which. is. consistent.
with.the.cylindrical.shape.of.the.embryo.being.the.major.cause.of.the.aberrations. . Figure.15.25c(i) . shows.
the.image.reconstructed.from.the.calculated.wavefront,.conirming.the.accuracy.of.the.calculation..If.
the.phase.of.the.wavefront.is.corrected,.the.peak.intensity.of.the.image.of.the.bead.would.increase.by.a.
factor.of.4.2,.and.the.shape.of.the.image.would.again.resemble.a.point.( Figure.15.25c(ii) ) ..he.amplitude.

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Iteration Iteration

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FIGuRE 15.23 (a). Maximum. intensity. of. the. measured. PSF. for. each. iteration. of. the. wavefront. optimization..
(b) Corresponding.Strehl.ratio.

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FIGuRE 15.24 Lateral.in-focus.cross.section.of.the.inal.PSF.from . Figure.15.22 . (circles)..Gaussian.it.to.the.data.
(gray.line)..heoretically.calculated.image.of.a.200.nm.bead.from.an.ideal.pupil.of.NA.1.285.(black.line).