PART THREE
Special Techniques in Cytology
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A
B
F ig . 3 3 .3
(A) This liquid cytocentrifuge
m o n o la y e r p r e p a r a tio n is
h o m o g e n e o u s ly fo c u s e d .
(B) In contrast,
s o m e c e lls o f th e g y n e c o lo g ic
s m e a r a re o u t o f fo c u s ,
as the slide comprises several focal planes.
2. z-sharp ening (extended focus im a g in g ): first, a z-stack is
acquired, th e n a softw are a lg o rith m extracts th e sharpest
areas fro m th e d iffe re n t focal planes and com b ines th e m
to a synthesized im age w ith a h ig h d ep th o f sharpness.
z-stacks m u ltip ly the storage requirem ents according to
the n um b er o f a d d itio na l focal planes th a t are scanned and
increase d o w nlo ad tim e w h e n vie w ing the slide on the In te rn e t.
z-sharpening is applicable fo r cells lyin g side b y side b u t in d if-
ferent focal planes o r uneven h isto lo g y sections. H ow ever, this
technique is unsuita b le fo r cell groups w ith overlapping hyper-
chrom atic nuclei, as nuclear details are com p letely effaced in the
com p ound image. B oth approaches sig n ific an tly increase to ta l
scanning tim e. As z-stacks m ore closely em ulate a real m ic ro -
scope, th is w ill be the m eth od o f choice fo r cytology smears. In
ord er to m in im iz e a d d itio na l scanning tim e and storage require-
m ents, the scanning o f m u ltip le focal planes m ay be restricted
to relevant cell groups instead o f the w h o le slide. Interfram e
com pression, a com pression procedure applied in videos, m ay
be a no the r p o ssib ility to reduce the storage requirem ents.
cover
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carrier
F ig . 3 3 .4 T h e d e p th o f fo c u s
comprises only part of the z-axis of a given
slide. (a) Slide areas above or below the depth of focus are out of focus if
only one focus plane is captured. (B) A conventional microscope allows
adjustment of the focus plane by moving the microscope focus knob. this
way, any area of the slide can be focused in the z-axis. this function of a
traditional microscope may be simulated by capturing all focal planes of a
given slide.
Applications
Teaching: Online Education
The advantages o f v irtu a l m icroscopy outw eig h the disadvan-
tages in educational applications b y fa r (Table 3 3.1) .
G o od microscopes are expensive, and cytologic preparations
fo r teaching are o fte n available in lim ite d num bers only. O ver
the years, the specimens fade, break, o r get lost. C ollections o f
v irtu a l slides propagated via the In te rn e t obviate the need fo r
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