NAME:


PRAC CLASS:














ANAT 2008: PRINCIPLES OF HISTOLOGY


Practical Book


2021











Discipline of Anatomy & Histology
University of Sydney NSW 2006


Course Coordinators and Editors: Dr Samson Dowland
sam.dowland@sydney.edu.au
Dr Katie Dixon
katie.dixon@sydney.edu.au


INTRODUCTION FOR ANAT2008 PRACTICAL CLASSES

REQUIREMENTS FOR PRAC CLASSES
No videoing or recording of the lectures or practical classes are permitted.
Recordings are made of the lectures and are available on Canvas. The practical sessions
are designed to allow sufficient time to take notes as well as the opportunity to ask the
tutors questions after the talk.

1. TEXTBOOKS
The textbook is HISTOLOGY: A TEXT AND ATLAS by W. Pawlina (Wolters Kluwer
Health, 8th edition 2019). It is recommended that this textbook is purchased prior to the
first practical class. The online version of this textbook is provided by the University Library
and a link is available on Canvas.

2. PRACTICAL BOOK
Students will build a record of their histological study throughout semester. Each topic will
have a set of practical notes, which can be combined into a practical book. The practical
notes will be available on Canvas for each topic and should be printed out prior to class. It
is the student’s responsibility to bring the copy of the practical notes to the appropriate
class. The practical notes are essential for each class.
The Histology exercises refer either to slides in the Discipline Collection (see below) or to
images in Pawlina, unless otherwise specified. The images required for practical exercises
are identified by a figure number followed by a page reference in brackets, e.g. Fig. 9.5
(279).
The Practical Book provides a permanent record of your observations and answers to the
questions and is a valuable resource for revision. The practical book does not get handed
in or marked.
IMPORTANT NOTE: The only students permitted to use practical books that have already
been worked in are repeating students who are updating their own books. In all other
cases, the use of other people’s books constitutes plagiarism and is therefore not allowed.

3. SLIDE COLLECTION
General Collection
This course uses sets of histological slides, which can be examined under the microscope.
The departmental slide collection has been prepared over many years from tissues
donated from many sources. It is your responsibility to care for these slides and ensure
they are kept in good condition for others to use. There are two labels on each slide. One
is a departmental code describing species, organ and preparative method. The other label
indicates the exact location of the slide. The colour of the label corresponds to the colour
of one of the spots on the box in which it is located. The numbers on the label indicate the
correct position of the slide in the slide box and the particular box in which the slide is kept.
For example, Slide Orange 3/128 is in box 128 with an orange spot. Please adhere to this
system.
Please check that no slide is left on your microscope.
A SECURITY DEVICE IS IN OPERATION IN THE TEACHING LABORATORIES. ANY
STUDENT ACTIVATING THIS DEVICE WITH UNIVERSITY PROPERTY WILL
AUTOMATICALLY BE REFERRED TO THE VICE-CHANCELLOR ON A PROCTORIAL
MATTER. NO EXCUSES WILL BE ACCEPTED.
NB. Students may not eat or drink in the laboratories.

4. ATTENDANCE AT PRACTICAL CLASSES
PLEASE NOTE: ATTENDANCE IN PRACTICAL CLASSES IS NOT MANDATORY IN
2021 DUE TO THE CONTINUING COVID-19 SITUATION. IF YOU FEEL UNWELL,
PLEASE DO NOT COME TO CLASS. There will be opportunities to catch up on any work
that is missed.
However, it is strongly encouraged that you attend all practical classes and complete all
practical exercises. These classes are essential for developing a full understanding of the
course material and to perform sufficiently in the final exams.
It is a good idea to attempt any questions not requiring the class slide collection prior to the
prac class to optimise time in class using the slides. Throughout the semester there will be
time in some practical classes to complete unfinished work or begin revision.
Due to the large number of students in this course, the allocation of practical classes is
completed by the timetable unit. There is no changing of practical classes unless there is a
documented clash with another subject. In order to arrange this you need to contact the
timetable unit.

5. ARRANGEMENTS FOR REVISION
Due to the large number of students enrolled in ANAT2008, students will only be permitted
to attend their allocated practical class.
Unfortunately due to funding constraints and WHS considerations, histology laboratories
will not be opened outside of normal class hours.
Revision sessions will be run during stuvac, further details will be provided. Under no
circumstances will students be allowed to attend sessions other than those allocated to
them.
There is ample time during semester to complete the exercises in the practical book. If a
practical session is missed, it is advised that these exercises be completed upon returning
to class. Tutors will be able to answer questions related to work in previous weeks.

6. DEPARTMENTAL NOTICES ON CANVAS
Information pertaining to this course is available via Canvas. It is the responsibility of the
individual students to check this regularly and to read carefully the information displayed.
Communication will also be made via university email.

EXAMINATIONS
NB: any changes to the following information will be displayed on Canvas. The
current method of examination is as follows:
• In-semester online theory quizzes will be completed on Canvas
(10mins and 5% each) and will be worth 15% of the final marks in
total.
• The mid-semester prac quiz (30mins) will include both theory and prac
questions and will be worth 15% of the final marks.
• The final theory examination (60mins, plus 10mins reading time) will be
worth 35% of the final marks.
• The final practical examination (60mins, plus 10mins reading time)
will be worth 35% of the final marks.
• Students must pass both theory and practical examinations to pass
the course overall.
• Marks allocated to each section of the course (e.g. cytology, etc.)
reflect the approximate time spent on that section relative to the
course as a whole.

Assessment
Name
Assessment
Category
Assessment
Type
Exam/Quiz
Type
Individual/
Group
Length/
Duration
Weight
Online
Theory
Quizzes
Quiz In‐Semester
Quiz
Canvas
Quiz
Individual 10mins each 5% each
(15% total)
Mid‐
Semester
Prac Quiz
Quiz In‐Semester
Quiz
MCQ
questions
Short
answer
Individual 30mins 15%
Final Exam ‐
Theory
Exam Final Exam MCQ
questions
Individual 60mins 35%
Final Exam ‐
Prac
Exam Final Exam Short
answer
Individual 60mins 35%

1. Theory Examinations

• The theory questions consist solely of multiple choice (MCQ) type
questions and approximately half a minute is allowed per question.
• Only one answer applies to each question and there is no negative marking.

• The final theory exam is supervised by the Registrar’s Office



2. Practical Examinations

• Students will be expected to recognise and answer questions on structures
in slides and in electron micrographs. Diagrams and other illustrative material may
also be included in the practical examination.
• The number of marks per question is shown on the examination paper and as a
rough guide, one mark is equivalent to one minute of examination time. Each section of
the examination is usually marked by the lecturer concerned.
• Students should ensure that they are fully conversant with the operation of the light
microscope as no assistance with setting up and use of the microscope will be given
during practical examinations.
• Please check Canvas closer to the examinations period for location of the practical
examination.
• There may be several sessions of practical exams to accommodate the large
number of students in this course. Please allow at least 3 hours from the start of the exam
sessions to allow for waiting time.
• Note that these exams could be at any time during the examination period and as
such please delay any travel until after the examination period. No special consideration
will be given under these circumstances.

1. Examination Results and Additional Information
• Students will be advised of their formal results in the period after the semester
exams.
• In cases of Special Consideration a replacement examination may be granted.
These are run by the faculty. The replacement practical exam will involve the use of slides
and microscopes as well as diagrams and illustrative material. The replacement theory
exam will involve M/C questions, short and long answer questions. If you are awarded a
replacement examination, then you must attend this alternate date. The results from the
original examination (for which Special Consideration was sought) will not be recorded.
• In the unlikely event of a second replacement exam, this will occur at a date/time to
be advised and is an oral exam in the presence of at least 2 academic members of staff.
Practical and theory questions will be asked from illustrative material and/or slides (which
will need to be viewed using a microscope).
• The further testing, if any, will be as soon as possible after the end of semester
exams and will be confined to one session only.
• In the case of replacement exams the course coordinators will contact students by
email to provide information on the format, location and other details of this further testing.

IMPORTANT NOTE
There is a strict examination cheating policy enforced by The University of Sydney (see
website for details). No cheating in any assessment will be tolerated and may lead to
expulsion from the course and disciplinary action taken by the university. In the past,
several incidences have led to disciplinary action.
INSTRUCTIONS GIVEN TO STUDENTS ON FRONT PAGE OF THE THEORY
EXAMINATION BOOKLET

ALL QUESTIONS TO BE ATTEMPTED. ALL ANSWERS ARE OF EQUAL VALUE
This paper contains xxx questions of the Multiple Choice type.
Each question has five possible answers – a to e. Each question has only ONE correct
answer. Note that some sets of answers are used for multiple questions. Each question
should be treated independently.

The answers should be entered directly onto the answer sheet. Answers written in the
question booklet will not be marked or transcribed to an answer sheet for machine
marking.

When filling in the answer sheet always check the question number before filling in
the circle. The circles should be blocked in BOLDLY and completely with a 2B lead
pencil. Any erasures must be complete. Incomplete erasures (i.e. pencil smudges) may be
detected by the marking sensor and scored as wrong answers.







POINTERS FOR PRACTICAL EXAMINATIONS,
(Or how not to make silly mistakes!)

Please note that slides you are asked to identify in a practical examination may not be the
same slides as those looked at in class. They may be from different species, stained
differently, or cut in a different direction. However, they will be no more difficult to interpret
than the class slides if you know your work. They are not labelled in the same way as your
class slide collection.
The following suggestions should help students during the final exam and in the practical
quizzes.

1. Check that the number on the slide is the same as the slide
number in the question, and that you have read the
number the right way up.
The example shown is Slide 61, not Slide 19.
Also ensure that the slide is the correct way up, that is, the
coverslip is on the top.



2. Look at the section with the naked eye to see its overall topography. Some tissues
and organs can be identified this way although, of course, you must check with the
microscope.

3. Quickly look at the entire section under low power to check whether there is one or
more than one organ on the slide, and to check for distinguishing features.

4. Know the definitive features that distinguish one organ from another.

5. Do not extrapolate beyond what is on the slide.

6. Use your common sense when you look at the section. Hair follicles are not found in
the trachea, or the uterus, or the skin of a lizard. Sperm are not found in sweat
glands.
Number
Cover slip
61
7. Answer questions concisely, usually with one or two words or a short descriptive
phrase. A long rigmarole will almost certainly introduce errors that could cost you
marks.

8. Give only one answer to each question, e.g. do not identify a tissue as “cartilage or
bone” and the cells within it as “chondrocytes or osteocytes”. Even if one set of
alternatives is correct, you will get no marks because you could not decide which
was the right answer.

9. Tissues are components of organs, e.g. the trachea (an organ) contains several
tissues including epithelial lining, cartilage, loose connective tissue, glandular
tissue, etc. If a question refers to one of these tissues specifically, so too must your
answer.

10. A hollow organ such as the trachea has an epithelial lining that forms its inner
surface. An outer surface has a covering. For example, a bone is covered by
periosteum and lined by endosteum.

However, a block of tissue may have been taken from the wall of a hollow organ: a
section from this block of tissue will be covered on one side by the epithelium that
lines the organ as a whole. (Note also that some text books etc. use the term
“lining” incorrectly to denote an outer covering.) We do our best to construct
questions so that you will have no doubt about their meaning.

11. When referring to connective tissues, distinguish between the cells and the non-
cellular (or acellular) matrix in which the cells are embedded. Both the cells and the
matrix constitute the tissue. When asked for the components of a matrix, mention
ground substance and fibrous component etc. but not cells or blood vessels.

12. Take care with planes of section. In some courses questions may be asked about
planes of section specifically, but all students of Histology should understand them.
Planes of section are also a matter of common sense, e.g. a tubular object cannot
have an equatorial plane of section because it has no equator. On the other hand,
an equatorial section is probably the one clearly identifiable plane of section through
a red blood cell.

13. With all questions, in theory as well as practical examinations, read all parts of the
question very carefully, paying particular attention to “words that describe”. e.g.
cells in the granular cell layer of the epidermis are packed with keratohyalin
granules, not melanin granules.

EXAMPLE OF PRACTICAL EXAMINATION QUESTIONS
Question 1 (5 marks)
Identify labels on the diagram.

Question 2 (5 marks)
Identify labels on the micrograph

https://www.shutterstock.com/image-photo/light-micrograph-crosssectioned-muscular-artery-showing-223918411
Question 3 (2 marks)
Identify the vessel in question 2, give 1 reason for your answer
A
B
C D
E
ANSWERS TO PRACTICAL QUESTIONS

Question 1 (5 marks)
A – lumen
B – secretory granules
C – rough endoplasmic reticulum
D – basal lamina (not basement membrane)
E – golgi
Question 2 (5 marks)
A – red blood cells
B – tunica intima (endothelial cells)
C – tunica media
D – tunica adventitia
E – internal elastic lamina
Question 3 (2 marks)
Medium muscular artery
Reason: mainly smooth muscle cells in tunica media

USE OF THE LIGHT MICROSCOPE

1. RESOLUTION AND THE OBJECTIVE LENS

The main function of the microscope is to resolve fine detail within the specimen. This
detail then has to be magnified sufficiently to provide a final image of a size which can be
seen by the human eye. The resolving power of the microscope is described by the
following equation:

dmin = λ 1
2NA

dmin is the minimum distance between two object points which can be seen as two
separate images.
λ is the wavelength of the light used (average of 500 nm for white light)
NA = n sin α
NA is numerical aperture
n is the refractive index of the medium between the slide and the objective lens
(in practice, either air or immersion oil)
α is half the intake angle of the objective lens in use







The maximum value for α is 72 which means that the maximum sin α is 0.95. The largest
NA which can be attained with a dry lens is therefore 0.95 (nair = 1).
The NA can be increased by putting oil between the slide and the objective lens. The
immersion oil is chosen so it has a refractive index equal to that of glass i.e., 1.5.
Therefore the maximum NA for the light microscope is 1.5 x 0.95 =1.4.
A small dmin means that very fine detail in the specimen can be resolved and this is
achieved when λ is small and NA is large. In practice λ is fixed since white light is usually
used but dmin can be varied by changing the NA.
The class microscopes are fitted with four objective lenses each with a different NA. They
range from low resolution, wide field of view 4X objective lens which has an NA of 0.1 to
the high resolution 100X objective lens which has an NA of 1.25.
NB. The NA and magnification are engraved on the barrel of each objective lens.

2. THE CONDENSER LENS

The function of the condenser lens is firstly to focus the light onto the specimen and
secondly, by means of the condenser iris diaphragm, to match the NA of the condenser to
the NA of the objective lens in use. This latter step is important because it can affect both
resolution and contrast as shown in the following diagrams:









(a) Setting of condenser iris is correct – the angle of the cone of light leaving the
specimen is equal to the intake angle of the objective lens.

(b) & (c) Setting of condenser iris is incorrect.

In (b) the angle of the cone of light is less than the intake angle of the objective lens which
reduces the effective NA of the objective lens and thus reduces resolution.
In (c) the angle of the cone of light is greater than the intake of the objective lens. This
results in a condition called “glare” in which excess light enters the lens. Glare causes
reduced contrast in the final image.
3. OCULARS
The role of the oculars is to further magnify the primary image formed by the objective
lens.

4. SETTING UP THE MICROSCOPE

a) Focus onto specimen using the 4X objective.
b) With the condenser iris fully open place a pencil point over the field lens of the light
source (on the base of the microscope). Rack the condenser up and down until a sharply
focused image of the pencil point is seen in the field of view. At this level light emerging
from the condenser is focused onto the specimen plane.
c) Remove an ocular and adjust the condenser iris to 7/8 of the back focal plane of the
objective lens. This procedure matches the NA of the condenser to the NA of the objective
lens.
d) When changing objectives the condenser iris should always be readjusted to 7/8 as
outlined in (c).

5. SOME PRACTICAL PROBLEMS THAT MAY BE ENCOUNTERED WHEN USING
THE LIGHT MICROSCOPE

After some experience students should be able to provide a reason and remedy for each
of the following problems.
(a) Problem: Inability to view field in both oculars.

Reason: Distance between oculars (interpupillary distance) incorrect. If difficulties are
still encountered after this distance is adjusted it may help if you move your head back a
little from the tops of the oculars.

(b) Problem: Dark lines or spots in field or diffraction boundaries around structures.
Reason: Condenser iris shut down too far.

(c) Problem: Inability to obtain sufficient light intensity.
Reason: Condenser lens racked down too far or condenser iris shut down too far.
(d) Problem: Poor contrast on 4X, 10X and 40X objectives.
Reason: Condenser iris open too far or transformer too high.

(e) Problem: Poor contrast or indistinct image on 100X oil immersion objective.
Reason: No oil or oil not making contact with objectives.

(f) Problem: Inability to focus microscope on high power objectives
(40X, 100X)
Reason: Slide upside down.

(g) Problem: Blurred image on 40X objective.
Reason: Oil on 40X objective or oil on slide.

(h) Problem: Field observed in one eye is out of focus.
Reason: Adjust focus of individual oculars.

(i) Problem: Part of the field of view is obscured.
Reason: Objectives not clicked correctly into place.

(j) Problem: Speckled or spotty image in one or both eyes. Spots do not move with
slide.
Reason: Dirt on oculars (twist oculars to see if spots move); or dirt or objective (move
objective lightly away from the clickstep position to see if spots move).
ANAT 2008 Student Feedback

Each year ANAT2008 is surveyed through the student questionnaires providing an
important opportunity for us to obtain student feedback. Suggestions from students have
resulted in modifications of the course, which have enhanced its delivery. Informal
feedback throughout the semester is also welcomed and can be made by emailing the
course co-ordinators.
Specific Feedback:
• Continual assessment and less dependency on one final exam
• Better explanation of the exam structure
• More detailed explanation of course assessment and value of each assessment
• More detailed objectives for lectures and practical exercises
• More time for revision
• Improve student feedback throughout the semester


The changes in response to the feedback:
• Multiple quizzes throughout semester collectively worth 30% of total mark
• A detailed list of objectives for each part of the course including the lecture and
practical components is at the front of the relevant chapter in the practical book.
• To pass ANAT2008 you must pass both the practical and theory components.
• Exam details are provided in the practical book as well as on Canvas and sent out
via email prior to the exam times.
• Trial theory exam questions (and answers) are at the end of each practical chapter.
These are the same format as the mid-semester exam and final theory exam.
• Trial prac exam questions (and answers) are at the start of the practical book.
These are the same format as can be expected in the mid-semester exam.
• A trial practical exam will be put onto Canvas prior to the revision session. Answers
will also be provided. This will be the same format as the final prac exam.



CHAPTER ONE – INTRODUCTION
HISTOLOGY
INTRODUCTION
The term ‘Histology’ literally refers to the study of the tissues of the body and the way in
which these tissues are organised to form the organs of the body (histos = tissue = woven
fabric; logia study or science of).
Cytology (= cell study) is obviously an integral part of histology and also (at the other end
of the scale) is microscopic anatomy or micro-anatomy of the organs within their organ
systems. An important branch of the science is the study of diseased or abnormal tissues
at the microscopic level – the science of histo-pathology.
In the earlier days of Histology the main emphasis was on morphological studies. This,
however, was largely a convenient division of research labour and expertise and functional
aspects were never far from the minds of histologists. The interdisciplinary nature of the
basic medical sciences becomes more evident as knowledge advances and you should
not be surprised if your teachers in Histology, Physiology, Biochemistry and Anatomy
seem occasionally to poach on one another’s fields.
Some Basic Concepts
The whole of the body is composed of cells, substances secreted by cells and fluids.
Cells, of course, are specialised both with respect to their behaviour towards neighbouring
cells and with respect to the type and amount of intracellular and extracellular material
they produce.
It is useful for descriptive purposes to recognise certain similarities and commonalities in
the properties of cells and their products and thus to group constellations of cells and cell
products to define a structural level higher than the cell and known as a tissue.
Epithelial tissue is one in which the cells closely cohere to one another and are separated
only by non-fibrillar intercellular material (intercellular cement). Such tissue is found
covering and lining body surfaces (e.g. skin, gut lining) and spaces (gland lumina). It may
also occur where a former connection with a surface has been lost (e.g. thyroid and
pituitary glands, thymus).
Nervous tissue consists of specialised cells called neurons – (specialised for electrical
conduction and the synthesis and release of transmitter substances) together with special
sheathing cells (neuroglial, Schwann and capsular cells). As in epithelial tissue the cells
cohere with one another.
Muscular tissue consists of cells which are specialised for contraction. The cells do not
cohere directly with one another except in some instances locally but are separated over
most or all of their extent by a layer of fibrillar material which is produced by the cells. This
material may be important in holding the cells in a coherent unit (e.g. smooth muscle).
Connective tissue is characterised by the presence of much intercellular material, material
which always contains a considerable fibrillar component. Included among the connective
tissues are fibrous connective tissue, bone, cartilage and the dental tissues dentine and
cementum.
Adipose tissue consists of cells specialised for the accumulation and release of lipid
materials (principally triglycerides). The cells are separated from one another by small
amounts of intercellular substance containing fibrillar material.
An organ (e.g. liver or kidney) is a functional and anatomical unit of body structure
composed of two or more tissues and usually having its own special nerve, blood and
lymphatic supply.
The functionally most ‘essential’ part of an organ (usually the epithelial or nervous tissue
cells performing the specific organ function) is sometimes referred to as the parenchyma
while the supporting tissue is called the stroma.
An organ system is an association of organs making up a major functional unit e.g. the
digestive or alimentary system of which the liver is one of the component organs or, say,
the urinary system of which the kidney is one component.

Methods of Studying
The present course in Histology combines light microscope and electron microscope
studies from the level of microscopic anatomy down to fine cell detail (Cytology). Structure
is related to function wherever possible. A significant improvement in the ability to discern
very fine structural detail has been made possible by the advent of the transmission
electron microscope. Studies using this technique are often referred to as ultrastructural
studies.
Other important forms of instrumentation will often be referred to in lectures. The following
list gives some idea of the range of equipment and techniques available. Light
microscopes (LM), including phase contrast, fluorescent, interference and polarising
microscopes, electron microscopes (EM) – either transmission (TEM) or scanning (SEM),
freeze-fracture techniques, histochemistry, autoradiography (using various isotopes),
microspectrophotometry, image analysis, cell culture techniques and so on. Consult
Pawlina, 8th Ed. 2019, Chapter 1 on “Methods”.

Some Basic Technical Considerations
In order to understand what you see when looking down the microscope or looking at a
micrograph it is necessary to be aware of a number of technical considerations.
1. Going from two to three dimensions
With conventional light microscopy although smears or spreads of whole cells may be
looked at (e.g. blood smears) it is more usual to look at thin slices of tissue (say 5-10mm
thick) and in the case of electron microscopy exceptionally thin or ultrathin slices of tissue
(say 50-100nm) are viewed. Because of this there is often initial difficulty with
interpretation – in other words it is not always easy to extrapolate from a thin 2D slice of
tissue back to the solid 3D object from which it came.
This concept may be further complicated by plane of section because many biological
structures are in the form of folded or convoluted tubes with perhaps a varying diameter
and maybe odd projections and the sections can be taken at different levels, different
angles etc.
2. Differences between LM & EM examination
Interpretation of EMs can be difficult by virtue of the very small volume of material being
examined and it should be fully appreciated that electron microscopy is an extension of
light microscopy, not a substitute for it.
For example, a piece of tissue as seen with the light microscope may contain thousands of
cells and many cells will be completely contained in the section (say section is 10µm) but
with EM sections, to reconstruct one 10μm diameter cell would require 100 relatively thick
sections.
The area of tissue which may be examined with the EM is always less than that possible
with LM. This is because it is necessary to have only very small pieces of tissue (1mm
cube) in order to be able to cut the very thin sections required (electrons are strongly
absorbed by tissues and sections only 0.5µm thick are virtually opaque). Also in order to
support so thin a section it must be mounted on a metal grid, the bars of which often give
an unobstructed viewing area only about 100µm square.
In volumetric terms we are here looking at say 1000µm3 of tissue as against, in an LM
section 1cm x 7μm, a volume of 7 x 108μm3.
3. Contrast
Contrast in microscopy is basically achieved by employing various staining techniques.
These procedures produce not only contrast of tissue relative to the background, but also
provide contrast between different elements within the tissues i.e. they differentiate
specific structures on the basis of their variable components.

Dimensions in Histology and Cytology
Based on one meter (m). 1mm (millimeter) = 10-3m
1μm (micrometer) = 10-6m; 1nm (nanometer) = 10-9m; 1pm (picometer) = 10-12m
A picogram (1pg) = is used for weight of cell components.