BIOLOGY 113 LABORATORY
Microscopes, Cells and Tissues
Objectives
• To learn the proper use and care of compound microscopes
• To learn staining techniques used in light microscopy
• To determine the magnification of the microscope
• To be able to identify the four major types of vertebrate tissues (epithelial,
connective, nervous and muscle).
• To understand how each type of tissue is organized and the distinguishing features
of each type of tissue.
• To associate tissue structure with its function for each tissue type.
Introduction
The ability to magnify specimens has been around since 1000 B.C. The first simple
compound microscope that utilized two lenses was not invented until the late 16th
Century. It was the invention and modification of this microscope that changed the way
scientists studied living organisms. It allowed scientists to study the structure of a living
organism and to discover numerous species that were not visible to the unaided eye.
Today there are numerous types of microscopes available to scientists that provide
greater magnification and superior detail (resolution). Besides magnifying and
resolving an object, the microscope also provides the contrast that is needed to
distinguish detail between adjacent objects. Microscopes used in most biology
laboratories magnify up to 1000X with a resolving power of 0.2 μM. The microscopes in
this laboratory are compound, light microscopes. The light is transmitted through the
specimen on the stage and through two lenses before it reaches the user.
These are expensive pieces of equipment. Please handle them carefully.
A. USING THE MICROSCOPE
There are some basic rules that you need to adhere to when using microscopes. These
are:
1. Always use two hands when moving your microscope. Use one hand to hold
the arm and the other hand should support the base. NOTE: The scopes are
heavy!
2. Use only LENS PAPER to clean the lenses. Do not use tissues, paper towels,
Kimwipes, your shirt, etc. to clean the scope. Even though these items may
feel soft, they can scratch the lenses.
3. The microscope must be on the lowest power objective lens:
a. when starting to use the microscope.
b. when you finish using the microscope. Before you return the microscope
to its numbered slot: 1) make sure the objective lens is on the lowest
power, 2) the power is off, 3) there is no slide on the stage and 4) the dust
cover is in place.
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4. NEVER use the course focus knob on high power; use ONLY the fine focus
knob. These microscopes are parfocal. This means that when the image is in
focus on one objective, the image will be in focus with the other objectives. You
may need to fine focus the image for sharpness.
Functional Parts of the Microscope:
Materials:
Microscope, lens paper, letter “e” slide
Procedure:
1. Remove the microscope from the scopes cabinet and return to your work area.
2. Make sure that you use both hands to support the microscope (arm and base).
3. Place the base securely on the lab bench with the arm towards you.
4. Identify the following parts of the microscope:
a. The ocular lens or eyepiece is a 10X lens which is at the upper end of the
tube. The scope that you are using is binocular; it has two eyepieces.
b. Revolving nosepiece: the objective lenses are attached below the
nosepiece. It allows the user to change the magnification.
c. Objective lenses: our scopes will either have 3 or 4 objective lenses
attached to a revolving nosepiece. The magnification is inscribed on each
lens. The powers that you will use are: scanning (4X), low (10X) and high
dry (40X). Most of our scopes also have an oil immersion objective lens
(100X). You will not use this lens in this lab.
d. Stage and stage clamp: the slide will rest on the stage and will be held in
place with the stage clamp. The moveable portion of the stage clamp
should only be touching one corner of the slide.
e. Iris diaphragm lever: on the front edge beneath the stage is a small lever
that is used to adjust the contrast by regulating the amount of light.
f. Condenser and adjustment knob: the condenser condenses the light rays
into a stronger beam. Use the adjustment knob located below and to the
side of the stage to increase or decrease the light intensity.
g. Coarse Adjustment Knob: On each side of the scope is a large knob use
to move the stage up and down to focus the image. This knob is to be
only used with the scanning and low power objective lenses.
h. Fine Adjustment Knob: Located by the coarse adjustment knob, this knob
allows for very small changes to the height of the stage. This knob is used
to increase the sharp focus of an image and is the only knob to be used
with high power.
i. Slide Movement Knobs: To one side of the stage, there are two black or
silver knobs that you will use to move the slide. One knob will move the
slide to the left and right. The other knob will move slide towards and
away from you. The scopes are parcentral. This means that when you
have the object centered in the field of view on a lower power and then
change to a higher power, the image will remain in the center of the field
of view.
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Introduction to the Human Tissues
Tissues are formed when cells with similar structure and embryonic origin are
aggregated together, performing a particular function. In this laboratory you will
examine the four major groups of tissues based upon their structure and function:
epithelial, connective, muscle and nervous tissues. The study of tissues, especially
the structure and arrangement, is called histology.
Today, we will examine humans from the tissue level of organization up. As you recall,
tissues are grouped to form organs which work together as systems.
Atomic  Molecular  Cellular  Tissue  Organs  Organ systems  Organism
A. EPITHELIAL TISSUE
Epithelial tissue form sheet like layer of cells with very close cell to cell contact.
They function in protection, absorption and excretion of materials for an organism.
Epithelial tissue is classified based upon 1) the number of layers and 2) the shape of the
cells on the free surface:
Simple epithelia are surface linings consisting of a single layer of cells. These are
usually found at places where selective diffusion, absorption, or secretion occurs.
Simple epithelia can range from squamous (flattened) to cubiodal to columnar in
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shape depending on their function. All epithelial cells secrete a non-living matrix called
the basement membrane. This is essentially a dense layer of extracellular material
that acts as a “glue” to hold cells together.
Simple squamous epithelium: a single layer of thin flattened cells, usually containing
a flattened nucleus. They line the lumens of the entire blood vascular system
(specifically called endothelium here), the renal corpuscles, and the lumenal surfaces
of the pleural and peritoneal membranes. In these areas, they function in diffusion of
materials. They also line the thoracic, abdominal and cardiac cavity (mesothelium).
• Observe dorsal view of the frog skin (may look hexagonal)
• Observe the cross section of a blood vessel and find the endothelium that lines
the lumen of the blood vessel.
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Simple cubiodal epithelium: a single layer of cube-shaped cells with a centrally
located nucleus. This epithelial tissue can be found in exocrine glands (salivary and
mucus), lining small ducts and tubules (kidney tubules). They function in secretion and
some diffusion. Other areas found: ovaries and pancreatic duct
• You may be looking at the kidney tubules; thus look for circular tubes composed
of simple cuboidal cells.
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Simple columnar epithelium: the height of these cells are great than the width and the
depth. All the cells touch the basement membrane and the nuclei are usually aligned in
the center or towards the bottom of the cells. Some cells are ciliated. These cells are
specialized in secretion and some absorption. Goblet cells are often associated with
columnar cells and secrete mucus. Are any of these cells ciliated?
Locations: nonciliated
ciliated
lines stomach & intestines
uterine tubes
some salivary glands
fallopian tubes
gall bladder
upper respiratory tract
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Psuedostratified columnar epithelium: all cells begin at a common basement
membrane but some cells do not reach the free surface. The nuclei in largest
dimension of cytoplasm; therefore, appear in two rows, thus the appearance of
stratification. Some may be ciliated. Typical locations include: eustachian tubes, upper
respiratory tract, epididymis, vas deferens. They are typically found in areas of high
abuse forming linings.
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Stratified squamous epithelium: composed of cells with many layers, cells flattened at
free surface but typically cuboidal or columnar at basement layer. They function in
protection and coverings, in areas with lots of abuse. Locations include: skin and lining
wet surfaces: mouth, esophagus, and vagina.
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Transitional epithelium: Highly modified, stratified squamous epithelium that lines only
the unrinary bladder, ureters and portions of the urethra. Cells of the basal layer appear
cuboidal or columnar. When the tissue is streched the cells flatten and look squamous.
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B. CONNECTIVE TISSUE
There are two types of connective tissue: loose, the most wide spread connective
tissue, and dense connective tissue. Connective tissue is composed of cells and
extracellular fibers imbedded in a ground substance (matrix). The ground substance
or matrix can be jelly-like, liquid or solid non-living cement of glue secreted by the cells.
There are three types of fibers that are typically found in the various types of connective
tissue. Elastic fibers are composed of elastin (protein) with a stretchy and flexible
characteristic. These fibers form thin wavy lines that can be stained purple, red or
brown in color. Collagen fibers are composed of collagen protein and found only in
animals. These fibers are characteristically thick and resist stretching. When stained,
they appear as thick wavy lines that are either pink or red. Reticular fibers are
inelastic and branching forming a network. These fibers are difficult to see without
specialized staining.
Cells that are found in connective tissues are named according to their function. Cells
called “blasts” are responsible for production a particular substance, “clasts” are
responsible for reabsorbing substances, and “cytes” are resting or mature cells that are
not producing or reabsorbing substances. For example, fibroblasts produce fibers.
Fibrocytes are mature fiber cells and fibroclasts are responsible for reabsorbing
(breaking down) fibers.
Loose Connective Tissue
Areolar connective tissue: this is the least specialized connective tissue in the adult
organism. This is essentially the “packing material” of the body. It is often found
anchoring blood vessels, nerves and body organs. Cells include fibroblasts,
macrophages, mast cells, adipose cells, and plasma cells. Also present are elastic
fibers and collagenous fibers. This tissue is found in the subcutaneous layer of skin; it
supports blood vessels, nerves and epithelia.
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Adipose tissue: the cells are the dominant feature with gigantic vacuoles for lipid
storage. Fibroblasts are present. Elastic & collagenous fibers are present but may not
be visible in sections. These fibers are laid down randomly in all directions. Adipose
tissue can be found nearly everywhere but especially around the kidneys, heart, eyes,
greater omentum, subcutaneous connective tissue.
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Dense Connective Tissue
White (dense) fibrous connective tissue: collagenous fibers are the dominant type of
fiber that’s arranged in wavy, parallel lines. Typical locations include tendons,
aponeuroses, cornea, most ligaments and periosteum. The cell that you’ll see in this
tissue is most-likely fibroblasts.
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Bone: bone is composed of collagen and mineralized calcium and phosphate salts.
Compact bone is composed of numerous Haversian systems. Within each Haverian
system there is a Haversian (osteon) canal that has an arteriole, venule, and nerve
supply. Osteoblasts, the bone forming cells that wall themselves in within the lacunae.
At that point they stop producing bone and are called osteocytes. The cells in lacunae
are interconnected by tiny branching channels called canaliculi. These channels allow
for the passage of nourishments to the cells from the blood supply. Nourishment can
also be passed from one osteon canal to another osteon canal via Volkmann’s canal.
The osteoblasts/osteocytes are arranged in concentric layers called lamellae (this is
similar to the growth rings of a tree). The function of bone is for mechanical support,
locomotion, protection and mineral salt reservoir.
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Hyaline cartilage: the most common type of cartilage (gristle). This cartilage contains
both elastic and collagenous fibers - neither is visible in light microscope prep. Within
the lacunae are the chondroblasts which produce the ground substance, cartilage. In
the outer portion of the cartilage are the chondrocytes. Locations include: articular
surface of bones, end of nose, fetal skeleton, trachea, costal cartilage
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Elastic cartilage: within the matrix, there are numerous elastic fibers (resilience &
flexibility). The lacunae are more densely clustered containing chondrocytes. Locations
include the external ear, auditory tube, and epiglottis.
Vascular Tissue: Blood and lymph are derived from loose connective tissue with
various cells called corpuscles (or formed elements) which are suspended in a fluid
matrix called plasma (ground substance).
Vascular tissue transports nutritive
substances, oxygen and hormones to tissues and carries away waste products.
Erythrocytes, red corpuscles or red blood cells (RBCs): humans typically have
approximately 4.5 - 5.0 x 106 RBCs/mm3. Mammalian RBCs are biconcave in shape
because there is no nucleus present. Avian and amphibian RBCs are nucleated. All
contain hemoglobin for oxygen and carbon dioxide transport.
Leukocytes, white corpuscles or white blood cells (WBCs): WBCs are
responsible for the defense. Some release chemicals while others are phagocytic.
Classification of WBCs are based on granules in the cytoplasm that stain.
Agranulocytes: without granules in the cytoplasm
o Lymphocytes: comprise 20-25% of the WBCs. They are small with a
large dark blue nucleus with thin crescent shaped rim of light blue
cytoplasm. These are involved in the specific immune response (T
lymphocytes). B lymphocytes are responsible for making antibodies
against foreign antigens.
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o Monocytes: comprise 3-8% of the WBCs. They have a large, kidney or
U-shaped nucleus. These are considerably larger than lymphocytes.
They are phagocytic on foreign bodies in loose connective tissue. They’re
called macrophages when they are actively phagocytizing foreign bodies.
Granulocytes: have granules that stain in the cytoplasm
o Neutrophils: are the most numerous of the WBCs (65-75%). They have
a nucleus with 3-5 interconnected lobes (typically 3 lobed). The granules
stain light pink-purple. They are phagocytic in connective tissue.
o Eosinophils: comprises 2-5% of WBCs. They have bilobed nucleus
that’s often difficult to see. The granules stain red-orange or what ever
color the red blood cells are. The function of eosinophils is still relatively
unknown. However, there is an increase in their numbers with parasitic
infections and allergic reactions. Some may even secrete histamine.
o Basophils: comprises less than 1% of WBCs. They have an S-shaped
nucleus that’s often obscured by the large, very dark purple granules. No
cytoplasm can be seen. If you do see blue cytoplasm, the cell is mostlikely a lymphocyte.
Basophils are involved in the non-specific
inflammatory response. They release histamine which causes redness
and swelling as a result of increased fluid movement into the area.
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MUSCLE TISSUE
Skeletal or striated muscle: You will observe two different sections of skeletal muscle,
longitudinal and cross. In the longitudinal section, the fibers are large, long and straight.
You should notice the long cells with striations and multiple peripheral nuclei. These
are among the largest cells in vertebrate body (1 fiber ~ 1 “cell”). Skeletal muscle cells
that are thin in are capable of rapid activity (i.e. extrinsic eye muscles) whereas cells
that are thicker, such as those found associated with the appendicular skeleton contract
more slowly but with greater force. These are also known as “voluntary” muscles of the
body because we have conscience control over them.
In cross section, each “circle” represents one fiber and gives the massive end view
of myofibrils. Note peripheral nuclei that may be present.
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Smooth/nonstriated muscle: Smooth muscles have more of a spindle shaped cell with
no striations. They are also mononucleated. These muscles are involuntary since we
do not have conscience control over these. They are found in organs of the body and
vessels.
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Cardiac muscle: Cardiac muscle is also striated; however, the cells are branched and
contain a single central nucleus. Between the cells are the intercalated disks with gap
junctions. Found in the heart.
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NERVOUS TISSUE
Nervous tissue is composed of neurons. The basic anatomy of neuron consists of a
cell body or soma that contains the nucleus. There may be numerous extensions
called dendrites around the soma which function in receiving electrical impulses from
various stimuli and transmitting this information to the soma. The longer axon then
carries the impulses away from the soma to a particular organ to elicit an affect. The
axon may or may not be myelinated. Myelinated axons are enclosed by concentric
rings of lipid layer that insulates the axon. The myelin sheath is produced by Schwann
cells. Between each myelin sheath are small gaps in which the axon is actually
exposed portion called nodes of Ranvier. As the nerve impulse is forced to “jump”
(salutatory conduction) from node to node down the axon, the rate of conduction is
increased. At the end of the axon, there are numerous axon termini which innervate
various structures. At the end of each axon terminus is the foot or terminal bouton.
The small gap between the terminal bouton and the innervated structure is called the
synapse. This is where the release of neurotransmitters takes place.
Giant multipolar neuron: In a smear from spinal cord, you will observe numerous
neurons that typically stain bluish-purple. You should be able to see the soma,
nucleus, nucleolus and dendrites.
Myelinated axon:
Schwann cells create the myelin sheaths that are composed of
lipids. Can you find the nodes of Ranvier?
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