1 - Cytology = introduction

• Cytology: study structure and function of the cell

• Histology: study tissues and microscopic structure of the organs
(general and organ histology)
• Embryology: study the fetal development of vertebrates

Origin and historical development

• 1st period (observations): 1st microscope = 1590 by Hans and
Sacharias Jansen. Improved version in 1610 by Galileo Galilei
• 1611-1628: Christoph Scheiner made real prototype of modern
microscope. Robert Hooke used similar microscope to examine cork –
discovery and use of the word “cell”
• Histology = Histos (tissue) + logia (science research). Term proposed by
Bichat (1771-1802)
• 2nd period: 1839 = Schleiden and Schwann independently postulated
cell theory
• Life exists only in the form of cells. All orga. composed of cells – it’s the
basic unit of all life functions, exchange, movement, reactivity and
continuity of information
• Each cell is derived from a cell
• 3rd period (study of the structure of cells and tissue): discoveries
made:
• 1825 – Purkinje = Cell nuclei
• 1852 – Virchow = every cell comes from a cell (Omnis cella e cullula)
• 1869 – Misher = nucleic acids (DNA, RNA)
• 1873 – Schneider and Fleming = mitosis
• 1874 – Bugli and 1875 van Benden = chromosomes
• 1885 – Boveri = centriole
• 1886 – Golgi = internal reticulum
• 1887-1889 – van Bender and Boveri = meiosis
• 1894 – Altman and Benda = mitochondria
• 1899-1908 – Emil Fisher = proteins and their amino acid organization
• 4th period (the development of molecular biology and cytology):
intro of new methods (cytochemistry, histochemistry,
immunocytochemistry, immunohistochemistry, electron microscopy, cell
and tissue culture and hybridization techniques)
• Accumulation of large amounts of scientific data = development of the
science

• Tissues = specialised to carry out different activities and specific

physiological functions in the body. E.g. excitability, conductivity,
 contractility, absorption and assimilation, secretion, excretion,
respiration, growth and reproduction.
• There are 4 basic tissues: epithelial, connective, muscular and nervous
• Epithelial: covers all free surfaces of the body –
protective and secretory function
• Connective: penetrates all other tissue –
supporting and protective function in the
construction of the organs
• Muscle: highly specialised for contractility – role
in movement of body parts
• Nerve: high specialization of cells for conduction
and contractility. Neurons connect the nervous system and body parts.

Principal methods in cytology and histology

• Light microscopy (routine, inverted,
phase contrast, fluorescence,
polarized and dark field)
• Electron microscopy (SEM, TEM and
fracture and freeze etching)
• Magnification: eyepiece x objective lens
• Inverted = light from top and objective
lense at the bottom. V. good for
observation of cell and tissue cultures,
including time lapse light microscopy of
living cells
• Phase contrast: based on the principle
that light changes its speed when
passing through cellular and extracellular
structures w/different refractive indices
(density) – image appears darker than
the surrounding background
• Fluorescence: structures/molecules of the cell are associated by
means of histochemical/immunohistochemical technique w/fluorescent
dyes and observed microscopically w/filters for a specific wave length.
• Polarizing: use polarized light. Commonly used to observe crystals.
Limited uses for biology as cells are fragile and they might die. Also
mostly colourless.
• Transmission EM: beam of electrons is transmitted through an ultra-
thin specimen, interacting with the specimen as it passes through it.
• Scanning EM: a beam of electrons is scanned over a specimen to
produce a magnified image of an object. The electrons interact with
atoms in the sample, producing various signals that contain information
about the sample's surface topography and composition.
• Freeze fracture: a specimen is frozen rapidly and cracked on a plane
through the tissue. This fracture occurs along weak portions of the
tissue such as membranes or surfaces of organelles. After cleaving,
both surfaces are shadowed with a platinum film. This coating
produces a replica of the surfaces.
• Can examine living cells and their tissues (thin material –
omentum/pleura, biological fluids and their cells and cell and tissue
 cultures) or post-mortem tissue sample or biopsy material to prepare a
permanent histological slide

Preparation of a permanent histological slide

• Coagulative fixatives (ethanol, matanol, HSI, chromic acid) fix material
for light microscopy. Precipitates (coagulate) and denatures protein.
Unsuitable for EM
• Non-coagulative fixes (formaldehyde, glutaraldehyde) – form inter and
intramolecular changes of the solution from solution to gel.
• Formaldehyde = quickly penetrates tissue but removed easily w/few
washes
• Glutaraldehyde = slowly penetrates but permanently binds to proteins.
Cell membrane is partially permeable and osmotically active
• Staining: most usable dyes are soluble in water therefore before
colouring must remove paraffin from the tissues by performing
rehydration
• Protocol Hematoxylin/Eosin stain = after rehydration place in
hematoxilin for 20-40 mins. Lavage 1-5 mins. Differentiation in ethanol
containing 1% HCl for 5 sec – removes excess paint and stains nuclei.
Wash in water. Stain in eosin solution for 10 min. Lavage. Dehydrate
and paraffin embedding.
• Preparation for EM (postfixation): osmium tetraoxide (OsO4) =
secondary fixative as it associates w/lipids as it’s believed that the
unsaturated bonds of the FA are oxidized by it and is materialized. It is
an electronically dense black substance that strengthens and increases
the contrast of the sample.