The document provides information on electron microscopy. It discusses the basic components and operating principles of transmission electron microscopes and scanning electron microscopes. Key points include: TEMs use electromagnetic lenses to focus electrons into an image, while SEMs scan specimen surfaces with a focused electron beam to produce topographical images. Both require specimens to be prepared through fixation, dehydration, embedding and sectioning to withstand the vacuum conditions. Contrast in electron micrographs is obtained through interactions between electrons and the specimen.

1. ELECTRON
MICROSCOPY
-Dr Ganga H

2. • Introduction
• Transmission EM
• Scanning EM
• Preparation of tissue for EM
• Diagnostic applications

3. Introduction and History
• The word microscope is derived
from the Greek mikros (small) and
skopeo (look at).
• The light microscope -developed
from the Galilean telescope during
the 17th century.
• Dutchman Antony van
Leeuwenhoek (1632-1723) –
simple microscope.
• Discovered protozoa,
spermatozoa, and bacteria, and
classified red blood cells by shape.

4. • The limiting factor in Van
Leeuwenhoek’s microscope was the
single convex lens.
• Addition of another lens could
magnify the image produced by the
first lens.
• This compound microscope –
consisting of an objective lens and an
eyepiece, a mirror or a source of light
and a specimen table for holding and
positioning the specimen.

5. Resolution of the Human Eye
• In sufficient light, the unaided human eye can
distinguish two points 0.2 mm apart.
• A lens or an assembly of lenses (a microscope) can be
used to magnify this distance and enable the eye to
see points even closer together than 0.2 mm.

6. Types of microscopes
• Three basic types:
• Optical, charged particle (electron and ion), or scanning
probe.
• Electron and ion microscopes, use a beam of charged
particles, and electromagnetic or electrostatic lenses
• They can see features as small a tenth of a nanometer,
such as individual atoms.
• Scanning probe microscopes use a physical probe (a very
small, very sharp needle) which scan over the sample in
contact or near-contact with the surface.
• These instruments are capable of atomic scale resolution

7. • A modern light microscope - 1000x .
• The resolving power of the microscope limited by
the wavelength of the light used for illumination.
• Using light with a shorter wavelength--a small
improvement.
• Using oil --- small improvement, but all together
only brought the resolving power of the
microscope to just under 100 nm.

8. • In the 1920- accelerated electrons behave in
vacuum much like light.
• They travel in straight lines and have wavelike
properties.
• Wavelength is about 1,00,000 times shorter
than that of visible light.
• Electric and magnetic fields are used to shape
the paths followed by electrons

9. 1931-Ernst Ruska at the University of Berlin built the
first transmission electron microscope (TEM)
1986- awarded the Nobel Prize for Physics

10. • The first electron microscope used two magnetic
lenses, and three years later he added a third lens
and demonstrated a resolution of 100 nm, twice
as good as that of the light microscope.
• Today, electron microscopes have reached
resolutions of better than 0.05 nm, more than
4000 times better than a typical light microscope
and 4,000,000 times better than the unaided eye.

12. Resolution of a microscope
• Wavelength of the illumination source ( λ )
• The numerical aperture of the lens (N.A.)
Limit of resolution = 0.61 λ/N.A.
• The maximum value of N.A. for light microscope is approx. 1.4.
therefore, that even the short blue light ( λ = 436 nm) of the
visible spectrum will yield a resolution of only 190 nm.
• The wavelength of an electron beam is about 100,000 times
less than that of visible light and hence the resolution of an
electron microscope is far superior to that of the light
microscope.

14. The Electron
• An atom is made up of three kinds of particles – protons,
neutrons, and electrons.
• The electrons, which are about 1800 times lighter than
the nuclear particles, occupy distinct orbits, each of
which can accommodate a fixed maximum number of
electrons.
• When electrons are liberated from the atom-they
behave like light.

15. Types of electron microscope
1. Transmission
electron
microscopy :
2. Scanning
electron
microscopy:

17. SEM

18. Normal cell under EM

19. Transmission Electron Microscope
Principle
• TEM is the direct counterpart of Light
microscope
• Involves passage of high velocity electron
beam through specimen, thin enough to
transmit 50% of the electrons
• Transmitted electrons – focused by lens
systems to form a 2 dimensional magnified
image

20. Analogy between LM & TEM
• Arrangement & function of their components
1. Illuminating system – source & condensor
2. Imaging system – lenses to produce
magnified image – objective & projector
3. Image translating system – Final image is
viewed

21. LM EM

22. THE LIGHT MICROSCOPE v THE ELECTRON MICROSCOPE
FEATURE LIGHT MICROSCOPE ELECTRON MICROSCOPE
Electromagnetic Visible light Electrons
spectrum used 390nm (red) – 760nm app. 4nm
Maximum app. 200nm or 0.14nm
resolving power 0.2micron Fine detail
Maximum
magnification x1000 – x1500 X 5,00,000
Radiation Tungsten or quartz High voltage (50kV)
source halogen lamp tungsten filament
Lenses Glass Magnets
Interior Air-filled Vacuum
Rigidly fixed, adjust
Focus Lens is movable
lens currents
© 2007 Paul Billiet ODWS

23. THE LIGHT MICROSCOPE v THE ELECTRON
MICROSCOPE
ELECTRON
FEATURE LIGHT MICROSCOPE
MICROSCOPE
Human eye (retina), Fluorescent screen,
Focussing screen photographic film
photographic film
Fixation formaldehyde Glutaraldehyde,OsO4
Embedding Wax Resin
Sectioning Microtome Ultramicrotome
slices - 20 000nm Slices - 50nm
Whole cells visible Parts of cells visible
Stains Water soluble dyes Heavy metals
Support Glass slide Copper grid
© 2007 Paul Billiet ODWS

24. • ELECTRON SOURCE
• ELECTROMAGNETIC LENS
SYSTEM
• SAMPLE HOLDER
• IMAGING SYSTEM.

25. Electron source
• The electron source consists of a
cathode and an anode.
• The cathode is a tungsten
filament which emits electrons
when being heated.
• A negative cap confines the
electrons into a loosely focused
beam.
• The beam is then accelerated
towards the specimen by the
positive anode.

26. Electromagnetic lens system
• The system allows electrons within a small
energy range to pass through, so the electrons
in the electron beam will have a well-defined
energy.
• 1. Magnetic Lens: Circular electro-
magnets capable of generating a precise
circular magnetic field. The field acts like an
optical lens to focus the electrons.
• 2. Aperture: A thin disk with a small (2-100
micrometers) circular through-hole. It is used
to restrict the electron beam and filter out
unwanted electrons before hitting the
specimen.

27. • TEM9.swf

28. The Vacuum System
• The electron beam must be generated in and traverse
through the microscope column under a high vacuum
condition.
• The presence of air molecules will result in the collision
and scattering of the electrons from their path.
• In the electron microscope the vacuum is maintained
by a series of highly efficient vacuum pumps.
• THE VACUUM FACTOR: Biological material must be
properly fixed and preserved

29. Sample holder
• The sample holder is a platform equipped with
a mechanical arm for holding the specimen
and controlling its position.

31. Imaging system
• The imaging system consists of another
electromagnetic lens system and a
screen.
• The electromagnetic lens - two lens, one
for refocusing the electrons after they
pass through the specimen, and the other
for enlarging the image and projecting it
onto the screen.
• The screen has a phosphorescent plate
which glows when being hit by electrons.

32. Image Formation in the TEM
• The basis of image formation in the TEM is the
scattering of electrons.
• The scattering results in a shadow on the viewing
screen or photographic film.
• Material with high atomic numbers will cause more
scattering and produce a deep shadow. Such material
is termed "electron dense" and has high image
contrast.
• Biological material has low electron density and is
known generally as "electron transparent". Hence, an
inherent low contrast image is formed.
• BIOLOGICAL MATERIAL must, therefore, be STAINED
with heavy metal salts.

33. Scanning electron microscopy

34. THE SCANNING ELECTRON
MICROSCOPE
• To directly visualise the surface topography of solid
unsectioned specimens.
• Probe scans the specimen in square raster pattern.
• The first scanning electron microscope (SEM) debuted in
1938 ( Von Ardenne) with the first commercial
instruments around 1965.
• Differs from TEM in construction & operational modes
• TEM – information is obtained from transmitted
electrons
• SEM – majority is obtained from secondary,
backscattered electrons & from X-rays.

37. Thin Specimen Interactions
• Incident electrons which are transmitted through the
thin specimen without any interaction occurring
inside the specimen- Unscattered Electrons.
• The transmission of unscattered electrons is inversely
proportional to the specimen thickness.
• Areas of the specimen that are thicker will have
fewer transmitted unscattered electrons and so will
appear darker, conversely the thinner areas will have
more transmitted and thus will appear lighter.

38. Elastic Interactions
• No energy is transferred from the electron to the
sample. The electron either passes without any
interaction or is scattered by electrostatic with the
positive potential inside the electron cloud.
• These signals are mainly exploited in TEM and
electron diffraction.

39. Inelastic scattering:
Primary electrons hit electrons of
the specimen atom
Energy is transferred from the
primary electron to the specimen
Emission of electrons and radiation

40. Electron specimen interaction

42. • After the impingement of the primary
electrons on the specimens, secondary
electrons as well as other forms of
radiation are emitted.
• But only the secondary electrons will be
collected by the signal detector.
• In the detector these electrons strike a
scintillator and the light produced is
converted to electric signals by a
photomultiplier.
• The electric signal is then amplified and
displayed on the cathode ray tube (CRT).

43. • In the SEM the electron beam is rapidly
scanned back and forth in an orderly pattern
across the specimen surface.
• It is a composite of many individual image
spots similar to the image formed on the TV
screen.
• The SEM has a specimen stage that allows the
specimen to move freely so that the surface of
the specimen can be viewed from all angles.

44. The focused electron beam is moved from one pixel to another.
At every pixel, the beam stays for a defined time and generates
a signal (e.g.secondary electrons) which are detected, amplified
and displayed on a computer screen

45. Image magnification in SEM
– A smaller area is scanned with the same number of
pixels.
– The scanned pixels are smaller

46. TEM vs SEM
TEM SEM
6 lenses – C1, C2, objective, 3 3 lenses – 2 condensor, 1
projector objective
High accelerating voltage - low accelerating voltage
penetration
Not complicated Specimen Stage – complicated
X & y axis X,Y,Z-axis, tilting, rotating

47. Contrast formation in TEM
Absorption of electrons
Scattering of electrons
Diffraction and phase contrast

48. Contrast formation in TEM
• Biological specimen consist of light elements:
Absorption contrast weak
Scattering contrast weak LOW CONTRAST
Phase contrast weak
• Contrast enhancement required:
– Treatment with heavy metals (Ur, Pb, Os)!
– Heavy metals attach differently to different
components

49. Thin section of alga stained with heavy
metals (Ur, Pb)

50. Contrast formation in SEM (using SE
and BSE)
• Different number of electrons from different
spots of the specimen
– Based on topography of the specimen
– Based on composition of the specimen

51. Uniform layer of heavy metal on specimen surface
Primary electron beam
Platinum

52. SCANNING TRANSMISSION ELECTRON MICROSCOPY
(STEM)
• This is a recent technological advance in the field of
Electron Microscopy.
• The beam of electrons scans the specimen, as it does in
scanning electron microscopy.
• However, it is the transmitted electrons that are
collected and amplified and form an image on a
cathode ray tube.
• The small spot size of the beam allows different areas
of the specimen to be discriminated and analyzed.
• A major use of STEM is in X-ray analysis which allows
the elemental composition of the specimen to be
mapped.

53. Specimen preparation for electron
microscopy
Steps include
 Specimen procurement
 Fixation
 Tissue processing and sectioning
 Staining

54. SPECIMEN PROCUREMENT
Tissue preserved in glutaraldehyde.
Tissue must be representative of the disease.
Areas that show - degeneration, necrosis,
haemorrhage must be avoided.

55. Drying of the surface must be avoided.
Tissue must be properly fixed.
The suitability of the tissue can be confirmed
by a frozen section or touch preparation.

56. • Fixation : most commonly used are osmium
tetroxide, glutaraldehyde and
paraformaldehyde.
• Dehydration : acetone or ascending
concentration of alcohol, 5-15 min in each
concentration.
• Use of dimethyoxypropane for rapid dehydration.
• Clearing agent: propylene oxide.

57. • Embedding media : methacrylate and epoxy
resins
• These medias infiltrate well and help in thin
sectioning
• Blocks are transferred to suitable capsule
containing fresh resin and these capsules are
transferred to incubator for polymerization.

58. Processing schedule
Fixation:
• Glutaraldehyde 2.5% at 4° C for 1-4 hrs.
• Wash in buffer.
Post fixation treatment:
• 1% osmium tetroxide at 4°C for 1 hr.
• Wash in water.

59. Dehydration
50% alcohol 5-15 min
70% alcohol 5-15 min
90% alcohol 5-15 min
Absolute alcohol 5-15 min
Absolute alcohol 5-15 min
Absolute alcohol 5-15 min
Clearing
Propylene oxide 15 min
Propylene oxide 15 min
Impregnation
Epoxy resin 45-60min
Polymerization at 60°C 24hrs

60.  When formalin fixed tissue used – area that is likely
to be fixed from outer surface to be chosen.
 Paraffin blocks-the corresponding light microscopic
section should be examined so that best portion of
the tissue can be mapped.
 However paraffin embedded tissue is never
satisfying for an electron microscopist because of
considerable distortion.

61.  Certain types specimens require special
processing unlike surgical specimens.
These include-
1. Percutaneous renal biopsies- 1-2mm pieces
from both the ends of the core are fixed to
ensure cortical glomeruli are represented in
tissue .

62. Aspirate directly expressed
into glutaraldehyde with
gentle agitation
and kept for fixation
2.FNAB
Filtration of the fixed
specimen through 20µm
mesh screen
Cells washed with pelleted
buffer and processed as for
solid tissue

63. 3.Bone marrow aspirate
Centifugation of Gentle Layering
Disk is gently
heparinized of fixative on the
transferred and
aspirate in buffy coat and
further processed
haematocrit tube fixation

64. 4.Core biopsies of bone
Challenging as decalcification causes severe
damage to cells.
Fixation in
glutaraldehyde
Soft marrow dislodged
with fine needle under
a dissecting microscope
Processed in routine
fashion

65. 5.Body fluids
Non • Centrifugation&
• Fixation in glutaraldehyde
hemorrhagic
fluids
Hemorrhagic • Erythrocytes removed
fluids with brief hemolysis.
• Rinsing in buffer and
fixation

66. TISSUE SECTIONING
Preparation of thick or semithin sections:
After the tissue has been embedded in plastic
resin
The blocks are embedded into sections at a
thickness of approximately 1µm.

67. And these are stained with methylene blue or
toulidine blue and
Examined to verify that blocks selected are
representative of the disease process

68. Thin sections are used for ultrastructural
study-50nm thickness.
These very thin sections are necessary-poor
penetrating properties of electron beam.
Ultramicrotomes are used for thin sectioning

69. STAINING
• Staining done using heavy metals such as
uranium and lead

70. Diagnostic applications
 As a rule the pathologist performing the EM
should come to presumptive diagnosis from
Clinical history and light microscopic findings
before performing the ultra structural studies.

71. Electron
microscopy
Ultrastructural
diagnosis of Ultrastructure of
non tumour tumors
biopsy

72. Non tumor biopsies Tumor diagnosis
• Epithelial tumors
• Diseases of kidney • Mesothelioma
• Metabolic storage diseases • Melanoma
• Respiratory tract biopsies • Hematopoietic and
• Skeletal muscle diseases lymphopoietic tumors
• Infectious agents • Soft-tissue tumors
• Cutaneous diseases • Central nervous system
• Peripheral nerve biopsies tumors
• Small round cell tumors

73. .

74. Renal biopsies
Aided in classification of renal disease in
particular & better understanding of the
pathogenesis of glomerular disease

75. a. Detailed study of glomerulus- epithelial cell,
endothelial cells, basement membrane &
mesangium.
b. Best method - to evaluate the thickness and the
structure of glomerular basement membrane.
c. Aids in identifying the exact location of immune-
complex deposits within glomerulus.

76. Renal B opsy D agnosi s U
i i sual l y
R equi r i ng El ect r on M cr oscopy
i
Minimal change nephropathy
Post-infectious glomerulonephritis
Membranoproliferative glomerulonephritis
Membranous nephropathy
Dense deposit disease
Diabetic nephropathy—early morphological changes (GBM thickening)
Fibrillary glomerulonephritis
Focal-segmental glomerulosclerosis—early recurrence in renal allograft

77. Algorithm of Interpretation of Ultrastructural Findings:
Discrete Immune-Type Electron-Dense Deposits Present
COMBINED
INTRAMEMBRANOUS SUBENDOTHELIAL,
(Usually Combined SUBEPITHELIAL,
SUBEPITHELIAL with Mesangial) SUBENDOTHELIAL MESANGIAL AND MESANGIAL
Membranous GN Dense deposit disease MPGN IgA Lupus (WHO
nephropathy classes III and IV)
Lupus (WHO class V) GN related to Lupus (WHO class Henoch- MPGN type III
endocarditis, deep- III and IV) Schönlein
seated abscesses purpura
Postinfectious GN Cryoglobulinemic Lupus (WHO GN related to
GN (microtubular class II) endocarditis,
substructure) C1q deep-seated
nephropathy abscesses
Rare other
forms of
mesangioprolif
erative GN

78. Algorithm of Interpretation of Ultrastructural Findings:
No Discrete Immune-Type Electron-Dense Deposits Present
Subendothelial
Fluffy Electron- Finely Granular Fibrillary/Microt
Normal GBM Diffusely Abnormal GBM Lucent Material Deposits ubular Deposits
Minimal change Diffuse thinning All forms of Monoclonal Amyloidosis,
disease, FSGS thrombotic immunoglobuli fibrillary GN,
Thin GBM disease, early microangiopathies, n deposition cryoglobulinemic
including malignantdisease GN, diabetic
hypertension glomeruloscleros
Alport syndrome is, collagen type
Diffuse thickening III
glomerulopathy
Diabetes, hypertension,
long-standing ischemia (also
wrinkling)
Diffuse lamellation /splitting
Alport syndrome

79. Minimal change disease
Extensive foot process effacement.

80. Post streptococcal glomerulonephritis
Subepithelial humps of IgG & C 3

81. Membranous glomerulopathy

82. Membranoproliferative
glomerulonephritis
TYPE 1 TYPE 2

83. Storage disorders
 Deposition - lipid and glycogen can be visualized
in biopsies of skin, brain, rectum, muscle, nerve,
spleen, lymph nodes , bone marrow, heart and
kidney.
 Gaucher’s disease- abnormal glucocerebroside
accumulation in reticuloendothelial cell of
liver,spleen,lymph nodes, and bone marrow.

84. Gaucher disease involving the bone marrow
A, Gaucher cells with abundant lipid-laden granular cytoplasm.
B, Electron micrograph of Gaucher cells with elongated distended lysosomes.

85. Ganglion cells in Tay-Sachs disease.
A, Under the light microscope, a large neuron has obvious lipid vacuolation.
B, A portion of a neuron under the electron microscope shows prominent lysosomes with
whorled configurations

86. Niemann -Pick disease
• Accumulation of
sphingomyelin in
lysosomes..
• Electron microscopy-
engorged secondary
lysosomes contain
membranous cytoplasmic
bodies resembling
concentric lamellated
myelin figures called
zebra bodies

87. Viral & other infections
• Body fluids, skin blister fluid, curetting from
warty skin lesions, surgically resected, PM
specimens
• Size 20-300nm
• Negative staining- 4% PTA

88. A, Adenovirus, an icosahedral nonenveloped DNA virus with fibers. B, Epstein-Barr virus, an
icosahedral enveloped DNA virus. C, Rotavirus, a nonenveloped, wheel-like, RNA virus. D,
Paramyxovirus, a spherical enveloped RNA virus. RNA is seen spilling out of the disrupted
virus

89. Electron microscopic picture of HSV

90. Whipple’s disease

91. Respiratory Tract Biopsies
 EM helps in studying several abnormalities of
ciliary structure.
 That is abnormalities in structure, number and
pattern of microtubules that compose the
axoneme of the cilium.

92. • Abnormal fine structure of cilia is seen in
ciliary dysfunction, such as immotile cilia
syndrome

93. Skeletal Muscle Biopsies
 Alterations that can be studied under EM are
relatively non specific.
 Inclusions within myofibrils - lysosomal and non
lysosomal storage disorders.*Fabry’s,Pompe’s+
 Congenital multicore disease - disaaray of myofibrils

94. Mitochondrial myopathy

95. Fabry’s Disease
Deficiency of alfa
galactosidase and
accumulation of
glycosphingolipids
Concentric
intracytoplasmic
inclusions

96. Ultrastructure of Tumors
• Electron microscopy is an useful adjuvant
techniques in the diagnosis and understanding
of neoplasms.
• Electron microscopy along with
immunohistochemistry is more helpful than
EM alone.

97. Indications
• Confirming the light microscopic diagnosis of a
neoplasm.
• Differentiating primary neoplasms from
metastatic neoplasms.
• Evaluating metastatic tumors of unknown
primary origin.
• Evaluating histologically undifferentiated
malignant neoplasms.

98. • Subtyping sarcomas
• Subtyping lymphomas and leukemias
• Evaluating neoplasms with unusual features
such as crystalloid inclusions.

99. Squamous cell carcinoma
Well differentiated squamous cell carcinoma
 Abundant cytokeratin filaments.
 Frequent desmosomes between cells.

100. Well differentiated squamous cell
carcinoma- frequent desmosomes

101. Poorly differentiated squamous cell carcinomas
Reduction in cytokeratin filaments
Reduction in desmosomes
Diminution in organelles
Loss of basal lamina

102. Adenocarcinoma
Microvilli – short, stubby, prominent
microfilament,& glycocalcyeal vesicles.
Few desmosomes and cytokeratin filaments.
Intracytoplasmic mucin or glycogen deposits
Tight junctional complexes

103. Tight junctional
complex

104. microvilli

105. Adenocarcinoma small intestine– demonstrates tight junctional
complex, mucin granules and luminal microvilli are typical .

106. Adenocarcinoma of colon- shows micrvilli, dense core rootlets below
and rounded glycocalyceal bodies in the villi

107. Ultastructure Of Mesotheliomas
EM helps in differentiating mesotheliomas
from adenocarcinoma.

108. Mesothelioma and Adenocarcinoma
• Mesotheliomas are characterized by having long,
narrow, branching microvilli with a length to
width ratio of around 10-16:1 ,-on free surfaces
of cells.
• By contrast, adenocarcinomas have short, stubby
microvilli with core rootlets.

109. Electron microphotograph of malignant mesothelioma-long thin non intestinal type of
microvilli devoid of glycocalyx and actinic rootlets.

110. Ultrastructure of Melanoma
Helpful in diagnosing melanomas -not express
anti-S-100 or the HMB-45 monoclonal
antibody.
In such cases identification of
premelanosomes or melanosomes hallmarks
the diagnosis of melanoma.

111. Melanosomes –cytoplasmic organelles
where melanin is produced.
 There are four stages in development.

113. Hematopoietic And Lymphocytic
Tumors

114. • Immunostaining remains the main stay.
• However EM is helpful –immunostaining is
equivocal or negative.

115. • Ultrastructural findings of leukemias and
lymphomas
1. Nuclear pockets
2. Absence of intercellular attachment
3. Lack of endoplasmic reticulum
4. Cytoplasm filled with free ribosomes
5. Sparse mitochondria
6. Lipid droplets in large cell lymphomas

116. 7. Immunoblastic transformation- the cisternae
of the endoplasmic reticulum become
abundant and organised.
8.Birbeck granules in Langerhans cells- are
endocytic organelles that transport antigens
from receptors on cell surface to interior to
fuse with saccules of the golgi complex
producing racket like configuration.

117. Burkitt's lymphoma
Numerous nuclear projections (np), polar aggregation of mitochondria (m),
sparse endoplasmic reticulum (er)

118. Histiocytosis X
• Mononuclear langerhans
cells with curved nucleus &
an abundant cytoplasm
• Birbeck granules in the
cytoplasm

119. Spindle cell tumors
• Fibrosarcomas- abundant rER, collagen formation
• Leiomyosarcomas – myofibrils & focal densities
• Spindle cell SCC- tonofibrils & occasional
desmosomes

120. Fibrosarcomas- abundant rER, collagen formation

121. Smooth muscle cell tumours- poorly developed ER , myofibrillary filaments attached to
focal densities

123. • Diffuse sheets not classified by other
means
• Myofilaments of skeletal muscle in
embryonal or alveolar rhabdomyosarcoma
• Lakes of glycogen in Ewing’s tumour
• Distinctive cytoplasmic processes in
neuroblastoma

124. Ewing’s sarcoma
Prominent lakes of glycogen

125. Embryonal RMS – cytoplasm showing haphazardly arranged abortive cross
striations

126. Neuroblastoma
Cytoplasmic
processes
wrapping around
a neuroblastoma
cell

127. Neuroendocrine tumours
• Neuro secretory vacuoles
in cytoplasm
• Spherical , ovoid
• Electron dense centre
surrounded by a clear
lucent halo enclosed in
distinct membrane.
• Carcinoids, APUD,
chemodactoma, medullary
ca thyroid

128. Central Nervous System
Neoplasms

129. • Classification of various cellular conformations of
the CNS is difficult when trying to distinguish glial
and neuronal elements using only light
microscopy and immunohistochemistry.
• Electron microscopy often plays a pivotal role in
diagnosis because it can provide accurate
diagnosis when immunohistochemical studies
are equivocal or negative.

130. Meningioma
• Long,interdigitating cellular processes
• Numerous cytoplasmic filaments
• Prominent desmosomes

131. Filgree pattern of the curving strands of attenuated cells- extracranial
meningioma

132. Disadvantages
1. EM is not economical- stable high voltage
supply, vaccum system etc
2. Findings unlikely to influence treatment, IHC n
LM together are confirmatory .
3. Tissue preparation is tough
4. Only a small proportion of neoplasm can be
studied
5. Misinterpretation of non- neoplastic elements
belonging to the tumor

133. Conclusion
Currently the use of EM is limited for the
expense and lack of surgical pathologists to
interpret EM findings .
Still it provides unique insight into the
structure of some tumors and renal
pathologies.
So better to use it selectively in study and
diagnosis of human diseases and research
areas and correlating the findings with LM
findings and IHC results.

134. Recent advances
• One of the latest developments in electron
microscopy is the environmental scanning
electron microscope (ESEM), which enables
soft, moist and/or electrically insulating
materials to be viewed without pre-treatment.

135. Hammar [2002] has succinctly
summarized the current
diagnostic status for IHC and EM:
“ There are no immunohistochemical
features that are absolutely specific
100% at this time in diagnosing a
neoplasm. There are a number of ultra
structural features of neoplasm that are
100% or nearly 100% specific in
diagnosing certain neoplasms."

136. References
• Theory and practices of histopathological
techniques, John D Bancroft, 4th edition;pg 585-
639.
• Robbins and Cotran, PATHOLOGIC BASIS OF
DISEASE, 8th edition.
• Cellular pathology technique, C.F.A. Culling, 4th
edition;pg 603-620.
• Various internet sources.

137. Thank
you