Biomedical Engineering Reference
In-Depth Information
performed  at. 50. μm. below. surface.. In. this. setting,. dominant. aberrations. in. this. rather. transparent.
sample.can.be.expected.to.originate.from.the.surface.proile.of.the.ish.head,.which.has.a.slightly.higher.
refractive.index.than.the.surrounding.agarose.gel..Without.wavefront.correction—that.is,.with.the.DM.
in.a.lat.shape—GFP.(green-luorescent-protein)-stained.neurons.and.their.processes.were.visible.(see.
Figure.14.9c ) .in.2PM,.but.resolution.was.notably.diminished.(as.indicated.by.the.lack.of.sharp.edges.in.
the.image)..Using.CGWS,.the.single-pass.aberrations.represented.by.the.lowest.28.Zernike.modes.could.
be.determined.(see. Figure.14.9b )..Applying.the.conjugate.of.this.optical.aberration.as.a.wavefront.bias.to.
the.DM.( Figure.14.9d ) .resulted.in.an.increase.of.69%.of.peak.luorescence,.along.with.a.visible.increase.
in. resolution.. Note. that. although. CGWS. is. in. principle. capable. of. single-step. wavefront. correction,.
Figure.14.9d. was.actually.a.result.of.ive.iterative.correction.cycles.involving.wavefront.measurement.
and.adjustment.of.the.DM..he.fact.that.further.improvement.was.possible.ater.the.irst.iteration.indi-
cates.that.the.response.of.the.DM.was.not.perfectly.incorporated.into.the.correction.algorithm.
14.5.3 Speckle Size and Multiple Scattering
As.the.CV.is.moved.deeper.into.a.strongly.scattering.sample.such.as.a.brain.tissue,.the.grain.size.of.the.
speckle.in.the.BFP.decreases..his.efect.can.be.seen.in.Figure.14.10,.where.CGWS.measurements.were.
performed.in.a.phantom.made.up.of.100.nm.beads.with.a.mean-free.path.(MFP).of.370.μm.down.to.ive.
MFPs..he.speckle.size.decreased.roughly.and.linearly.with.depth.
In.principle,.the.speckle.size.corresponds.to.the.inverse.of.the.lateral.extent.of.the.CV*.as.viewed.from.
the.detection.pupil..However,.the.lateral.extent.of.the.CV.should.not.depend.on.the.depth.inside.the.
scattering.sample,.at.least.not.when.single.scattering.is.dominant..his.implies.that.even.though.coher-
ence.gating.rejects.most.of.the.multiply.scattered.light,.its.contribution.and.thereby.the.efective.extent.
of.the.CV.increases.with.focus.depth..Since.this.multiply.scattered.light.passes.the.coherence.gate,.it.has.
the.same.optical.path.length.as.the.light.traveling.to.the.focal.region.and.exhibiting.one.scattering.event..
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Focus penetration depth within sample ( µ m)
FIGuRE 14.10 In.a.scattering.phantom.made.of.100.nm.beads.in.agarose.with.a.mean-free.path.of.370.μm,.the.
observed.speckle.size.decreases.with.focus.penetration.depth.
* . When.the.coherence.length.is.longer.than.the.Rayleigh.length,.the.lateral.extend.of.the.CV.can.be.much.larger.than.the.
difraction.limit.due.to.its.axial.extend.into.the.geometrical.part.of.the.beam.
 
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