Microbiology

Chapter 9: Biotechnology and Recombinant DNA

Biotechnology – the use of microorganisms, cells, or cell components to make a product

Genetic Engineering – manufacturing and manipulating genetic material in vitro;

inserting genes into cells; also called Recombinant DNA Technology

• gene from a vertebrate animal, including humans, can be inserted into the DNA of
a bacterium, or a gene from a virus into a yeast cell
o the recipient bacterium can then be made to express the gene, which may
code for a commercially useful product

Steps for Recombinant DNA Procedure:

1. Vector such as a plasmid is isolated
a. Plasmid – a small circular DNA molecule that replicates independently of
the chromosome
2. DNA is cleaved by an enzyme into fragments
3. gene fragment of interest is inserted into plasmid
4. plasmid is taken up by a cell such as a bacterium; plasmid multiplies
5. cells with gene of interest are cultured and then cloned
a. Clone – a population of cells arising from a single parent cell
6. Two possibilities:
a. copies of gene are harvested
b. bacterial cells make a protein product
i. copies of protein are harvested

Tools of Biotechnology:
Artificial selection – choosing one organism from a population to grow because of its
desirable traits

Site-directed mutagenesis – techniques used to modify a gene in a specific location to

produce the desired polypeptide

Restriction enzyme – an enzyme that cuts double-stranded DNA at specific sites

between nucleotides

• a restriction enzyme recognizes and cuts, or digests, only one particular sequence
of nucleotide bases in DNA, and it cuts this sequence in the same way each time
• restriction enzymes used in cloning experiments recognize four-, six- or eight-
base sequences

Methylates – addition of a methyl group (-CH3) to a molecule;

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 • bacterial DNA can be protected from digestion/enzyme cutting because the cell
methylates some of the cytosines in its DNA

Restriction Enzyme Process Steps:

1. restriction enzyme cuts double-stranded DNA at its particular recognition sites
2. these cutes produce a DNA fragment with sticky ends
3. when two such fragments of DNA cut by the same restriction enzyme come
together, they can join by base pairing
4. the joined fragments will usually form either a linear molecule or a circular one
5. the enzyme DNA ligase is used to unite the backbones of the two DNA
fragments, producing a molecule of recombinant DNA

DNA Ligase – joins the discontinuous fragments of the lagging strand

Vector – a plasmid or virus used in genetic engineering to insert genes into a cell; serve
as vehicles for the replication of desired DNA sequence

• a vector must be capable of replicating

• any DNA that is cloned in the vector will be replicated in the process

• when it is necessary to retrieve cells containing the vector, a marker gene

contained within the vector can often help make selection easy
o common selectable marker genes are for antibiotic resistance or for an
enzyme that carrier out an easily identifiable reaction
• plasmids are one of the primary vectors in use

viral DNA vectors – can usually accept much larger pieces of foreign DNA than
plasmids can

shuttle vectors – a plasmid that can exist in several different species; used in genetic
engineering

Polymerase Chain Reaction (PCR) – is a technique by which small samples of DNA

can be quickly amplified, that is, increased to quantities that are large enough for analysis

Steps:
1) incubate target DNA at 94 degrees Celsius for 1 minute to separate strands
(denaturation)
2) add primers, nucleotides (deoxynucleotides A, C, G, T) and DNA polymerase
3) primers attach to single-stranded DNA during incubation at 60 degrees Celsius for
1 minute
4) incubate at 72 degrees Celsius for one minute; during this time, two copies of
target DNA are formed

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 5) repeat the cycle of heating and cooling to make two more copies of target DNA

• 70 degrees Celsius polymerase activity

• annealing temperature (45 degrees and 70 degree Celsius variable)

Techniques of Genetic Engineering:

Inserting Foreign DNA in cells:
1) transformation – the process in which genes are transferred from one bacterium
to another as “naked” DNA in solution; DNA is precipitated with calcium
phosphate and applied to cells (used for bacterial and eukaryotic cells)
a. some lipid based chemicals also used
2) electroporation – a technique by which DNA is inserted into a cell using an
electric current; electrocuting cells in the presence of DNA
3) protoplast fusion – a method of joining two cells by first removing their cell
walls; is a way to isolate entire chromosomes; PEG (polyethylglycol) which
causes fusion of cell membrane; cells of different species can be fused
4) microinjection – the introduction of material, such as DNA, directly into a cell,
usually with a glass pipette; using a microscope and microneedle, DNA is injected
into cells
5) microprojectile gun – Tungsten bullets are coated with DNA and are shot into
plant cells

Introns – a region of a eukaryotic gene that does not code for a protein or mRNA

Exons – a region of a eukaryotic gene that encodes a protein

Reagents used in microbiology:

1) genomic library – contain natural copies of genes
a. restricted DNA is cloned into bacteriophage vectors; the phage infects
bacteria; multiplies each clone
b. each clone represents a “book” and all the clones represent a “library” or
“the human genome”
c. limitations: cannot really pull out or make a functional genes, due to
presence of introns; cannot make recombinant proteins
d. used for:
i. transcription
ii. Studying splicing
iii. Information about certain intronic DNA is very important, for
example fingerprinting
iv. Telomers – end of chromosome, important for studying aging
v. Mutation analysis of introns

2) cDNA libraries – contain copies of genes made from mRNA

a. circumvent introns

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 b. take mRNA (lacks introns) through isolation, invoke reverse transcription
resulting in complementary DNA (cDNA); than make a double-stranded
cDNA and than clone into bacteriophage vectors
c. from this process, genes lacking introns can be isolated and made
d. this library is used to make proteins

Synthetic DNA – under certain circumstances, “this” can be made in vitro with the help
of DNA synthesis machines
• in the 1960’s, it took Khorana 30 post_docs to make about 15 MER (base pair)
DNA outside a cell
• today, a high school student, with proper instruction, can make 20 MER (base
pair) DNA in about an hour
• can be used to make entire genes like insulin

Selecting a Clone:
• in cloning, it is necessary to select a particular cell that contains the specific gene
of interest
• this is difficult because out of millions of cells, only very few of those cells might
contain the desired gene

Two Methods for selecting a clone:

1) Blue-White Selection – using β-Galactosidase with X-Gal, bacteria turns blue;
however when the β-Galactosidase is disrupted , the bacteria turns white,
disruption caused by the fact that a gene was inserted into the β-Galactosidase
a. Plasmid DNA and foreign DNA are both cut with the same restriction
enzyme
b. Foreign DNA is inserted into the plasmid, where it inactivates the lacZ
gene
c. The recombinant plasmid is introduced into a bacterium, which becomes
ampicillin-resistant
d. All treated bacteria are spread on a nutrient agar plate containing
ampicillin and β-Galactosidase substrate, and incubated
e. White colonies that appear must contain foreign DNA
f. Blue colonies must not contain foreign DNA
• It is not a sensitive method and there can be background

2) Colony Hybridization – the identification of a colony containing a desired gene

by using a DNA probe that is complementary to that gene; highly selective
method which is highly sensitive
a. Create a master plate with colonies of bacteria containing cloned segments
of foreign genes
b. Make replica of master plate on nitrocellulose filter paper by placing the
filter paper on top of the master plate
c. Treat filter paper with detergent (SDS) to lyse bacteria

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 d. Treat filter paper with sodium hydroxide (NaOH) to separate DNA into
single strands
e. Add radioactively labeled probes
f. Probe will hybridize with desired gene from bacterial cells
g. Wash filter paper to remove unbound probe and expose filter paper to x-
Ray film
h. Developed film is compared with replica of master plate to identify
colonies containing gene of interest

Making a Gene Product:

• we have a gene
• we want to make recombinant protein
• utilize specialized vectors of plasmids from the cDNA library
• gene is transformed into bacteria from the promoter which is highly active in
bacteria
• RNA in bacteria is produced
• protein of interest in bacteria results

• using recombinant DNA techniques one can also make vaccines

o we cannot use attenuated HIV for vaccination
• this is called sub-unit vaccination

Human Genome Project – the goal of this project is to map the 35,000-70,000 genes in
human DNA and to sequence the entire genome, approximately 3 billion nucleotide pairs
• was started to map and sequence the entire human genome
o sequence – to formulate the exact order of nucleotides
• there are 3 billion bases in the human genome
• of those 3 billion, 10% are coding, 90% are noncoding
• Francis Collins – Human Genome Project director
o There are 20,000 to 30,000 genes

Scientific Applications of Microbiology:

• DNA sequencing – the process by which the nucleotide sequence of DNA is
determined
• Bioinformatics – the science of determining the function of genes through
computer-assisted analysis; computer assisted DNA Analysis
• Proteomics – study of proteins; the science of determining all of the proteins
expressed in a cell

DNA Fingerprinting – analysis of DNA by electrophoresis of restriction enzyme

fragments of the DNA; an important application for identifying an individual; parental
origin and even groups of people

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How is DNA Fingerprinting done?
• there are some areas of genome, mostly in introns, which are highly variable,
meaning it differs from individual to individual
• for example, there are some CA repeats (also called Variable Number Terminal
Repeats: VNTR)
• “CA” repeats can range from 4 to 80 depending on the individual
• Two methods for Fingerprinting:
o Southern Blotting – a technique that uses DNA probes to detect the
presence of specific DNA in restriction fragments separated by
electrophoresis
o PCR Amplification

Agricultural Applications:
• Ti Plasmid – an Agrobacterium plasmid carrying genes for tumor induction in
plants
o Crown Gall – a disease in tomatoes caused by a bacteria called
agrobacterium tumufaciens

Utilized Ti plasmid and goes and integrates into genome of tomato
cells

• herbicide resistance gene which was inserted into Crown Gall disease and through
the utilization of Ti plasmid, infect tomatoes, resulting in a resistant to herbicides
• put an anti-sense gene for polygactouranase  overripe

• anti-sense technique – is a technique for inhibiting gene expression at the RNA

level

Chapter 13: Viruses and Prions

• Mayer – showed a transmissible factor involved in Tobacco Mosaic Disease

• Iwanoski – showed that a filterable agent caused disease
• Wendel Stanley – isolated the first virus, derived from Yellow Fever

General Characteristics of a Virus:

• they are considered both living and nonliving
• Contain a single type of nucleic acid, either RNA or DNA as genome
• capsid (capsomers) – They have a protein coat called “this”
• multiply inside living cells by using the synthesizing machinery of the cell

Host Ranges:
• invertebrates
• vertebrates
• plants
• fungi

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 • bacteria

phages – viruses that infect bacteria are called “this”

bacteriophage – a phage can also be termed a “this”

Very large viruses; as large as small bacteria:

• Vaccinia Virus
• Ebola Virus

Viral Structure:
Virion – is a complete, fully developed, infectious viral particle composed of nucleic
acid and surrounded by a protein coat that protects it from the environment and it a
vehicle of transmission from one host cell to another

Major classification of a virus is by:

• shape of protein coat
• type of nucleic acid

capsid – the nucleic acid of a virus is surrounded by a protein coat called the “this”

capsomeres – each capsid is composed of protein subunits called “this”

envelope – in some viruses, the capsid is covered by a “this”, plasma membrane-like;

derived from most cells; usually consists of some combination of lipids, proteins, and
carbohydrates

spikes – depending on the virus, envelopes may or may not be covered by “this”, which
are carbohydrate-protein complexes that project from the surface of the envelope

nonenveloped capsids – viruses whose capsids are not covered by an envelope are
known as “this”
• the capsid of a nonenveloped virus protects the nucleic acid from nuclease
enzymes in biological fluids and promotes the virus’s attachment in to susceptible
host cells

General Morphology:
• helical – long cylindrical viral nucleic acid has “this” shape
• polyhedral – many-sided icosahedrons (20 triangular faces and 12 corners)
• Enveloped:
o Enveloped Helical - influenza
o Enveloped Polyhedral – herpes simplex
• complex viruses – some viruses, particularly bacterial viruses (phages), have
complicated structures and are called “this”

Taxonomy:

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 • Older classification of virus was based on the symptomatology disease

ICVT – international committee on taxonomy of viruses

ICVT groups viruses into families based on:

1) nucleic acid type
2) strategy for replication
3) morphology

virus – genus names end in “this”

viridae – family names end in “this”
ales – order names end in “this”

Example: For Human Herpes Virus 2:

Herpeviridae – the family name is “this”

Simplexvirus – the genus name is “this”

Viral Species – is group of virus sharing same genetic information and host range

Names of Viruses to know: p.382 - 383

1) parvoviridae
2) adenoviridae
3) poxviridae
4) herpesviridae
5) hepadnaviridae
6) picornaviradae
7) rhabdoviridae
8) filoviridae
9) paramyxoviridae
10) orthomyxoviridae
11) retroviridae
12) reoviridae

Isolation, Cultivation and Identification of Viruses:

• virus grow in most cells
• phages – most easy to grow are “these”
• phages can be stored in buffer at 4 degrees Celsius for months or can be frozen for
years at -80 degrees Celsius

Steps for growing viruses:

1) add phage to bacteria
2) phage infects bacteria resulting in phage/bacteria mix
3) add agar and place in plate

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 4) plaques – “this” forms, which are lysed bacteria where the virus grew and broke
out of the bacteria
Plaque Forming Units (pfu) – the concentration of viral suspensions measured by the
number of plaques are usually given in terms of “this”

• Phages Titer – are referred to as pfu – Plaque Forming Units

Growing Animal Viruses:

SIV – HIV virus in chimps
FIV – feline immune deficiency virus

• growing these animal viruses and studying these in a model similar to humans can
be used to help defeat human related viruses

embryonated eggs – injection of virus into eggs

cell cultures – most commonly used method to grow and propagate viruses

Life Cycle of a Phage:

Two Different Life Cycles:
• Lytic Cycle – ends with the bacteria lysing and the viruses escaping from the
lysed bacteria
a. Steps:
i. Attachment – phage attaches to host cell
ii. Penetration – phage penetrates host cell and injects its DNA
iii. Biosynthesis – Phage DNA directs synthesis of viral components
by the host cell
iv. Maturation – viral components are assembled into virions
v. Release – host cell lyses and new virions are released

• Lysogenic Cycle – bacteria remains alive, the virus forms a symbiotic

relationship with the bacteria
a. Steps:
i. Attachment – phage attaches to host cell
ii. Penetration – phage penetrates host cell and injects its DNA
iii. Viral DNA integration into bacterial genome
iv. Infected bacteria goes on replicating more infected bacteria

Vaccinations: p.510 - 511

• attenuated whole vaccines – use living but weakened virus
a. polio, MMR (measles, mumps, rubella (German measles)
• inactivated whole agent vaccines – virus are killed by formalin or phenol
a. rabies, influenza
• subunit vaccines – use only those antigenic fragments of a microorganism that
best stimulate an immune system response

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Prions – proteins that causes the neurological disease: Mad Cow Disease
• Stan Prusner – discovered prions
• there are 9 different forms of bovine spongiform encephalopathy

Normal Host Glycoprotein - PrPc

Diseased Host Glycoprotein – PrPsc - s = scrapie

Steps to Mad Cow Disease infection:

1) PrPC is located on chromosome 20 in humans and is secreted to the cell surface
2) PrPSc reacts with PrPC on the cell surface
3) PrPC converts to PrPSc
4) PrPSc is taken in by endocytosis and accumulates in lysosomes resulting in the
eventual formation of Mad Cow Disease

Viruses & Cancer:

• Several types of cancer are now known to be caused by viruses
• Sarcoma – cancer of connective tissue
• early in the century, chicken cancers were recognized
o caused by virus

• oncogenes – a gene that can bring about malignant/cancerous transformation

• oncogenic virus (oncovirus) – a virus that is capable producing tumors
• human oncoviruses:
o Epstein Barr Virus (EBV) – causes burkitts lymphoma and
nasopharyngeal lymphoma

90% of population in united states are carriers of EBV
o Hepatitis B – risk of liver cancer
o RNA Oncoviruses – HTLV-1 & 2 ATLs or Adult Cell Leukemia

Chapter 16: Nonspecific Defense of the Host

Nonspecific Resistance:
First Line of Defense:
• intact skin
• mucous membranes
• normal microbes in the GI Tract which inhibit growth of pathogens

Second Line of Defense:

• phagocytic white blood cells
o microphages
o neutrophiles
• inflammation

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 • fever
• antimicrobial substances

Third Line of Defense:

• specialized lymphocytes
o B & T Cells (White Blood Cells)
• Antibodies

Skin – present mechanical barriers against outside pathogens

• Epidermis (4-5 layer)
o Keratin – the top layer of epidermal cells is dead and contains a
protective protein called “this”, which causes:

dryness

Several layers are present

Skin Shedding
• Dermis (2 layers) – much thicker layer, containing blood vessels and lymphatic
vessels and other connective tissue
o Lymphatic vessels – are conduits for cells involved in immunity
o during inflammation, blood and lymphatic vessels are used for cells to
migrate to the site of injury where infection is thwarted

Mucous Membranes – are internal cavities, GI Tract, and Respiratory Tract are covered
in “this”s; present mechanical barriers against outside pathogens

goblet cells – mucous is secreted by specialized cells called “this”

Mucous – is made up of glycoproteins and it restricts growth and infection by pathogenic

microbes

Lacrimal apparatus – protects the eyes by producing tears and pathogenic microbes are
washed away or digested by an enzyme called lysozyme

Buccal Cavity – saliva is produced from three glands, contains lysozyme and amylase

Cilia – physically captures pathogens and also causes mucous to move

Chemical Factors:
Sebum – sebaceous glands of the skin produce an oily substance called “this”, which
inhibits growth of microbes

Perspiration – secreted by the sweat glands, helps maintain body temperature, eliminate
certain wastes and flushes microorganisms from the surface of the skin

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Lysozyme – perspiration also contains “this”, which is an enzyme capable of breaking
down cell walls of gram-positive bacteria and, to a lesser extent, gram-negative bacteria

Gastric Juices – acidic enzymes such as HCI (hydrochloric acid) produced by the glands
of the stomach; is sufficient in destroying bacteria and most bacterial toxins

Normal Microbiota and Non-specific Resistance:

Symbionts – in GI Tract, prevent growth of pathogens by altering pH and oxygen
availability

Blood is made up of:

• Erythrocytes – are red blood cells; make up most of the blood (4.8 to 5.4
million); involved in oxygen and carbon-dioxide transport
• Leukocytes – are white blood cells; make up a smaller amount of the blood
( 5,000 to 10,000)
o Granulocytes – white blood cells that owe their name to the presence of
large granules in their cytoplasm that can be seen under a light microscope
after staining

Neutrophiles – make up 60% to 70% of leukocytes - very highly
phagocytic in the first 5 minutes of infection

Basophiles – .5% to 1% - produce histamines

Eosinophiles – 2% to 4% - toxic protein production, some
phagocytosis

o Agranulocytes – while blood cells that also have granules in their

cytoplasm but they are not visible under the light microscope after staining

Monosites – 3% to 8%, when entering body tissue, they
activate/mature to form macrophages performing phagocytosis

Lymphocytes – 20% to 25%, involved in specific immunity
• Examples of Lymphocytes:
o B Cells – provide antibodies and grow to form
plasma cells
o T Cells – are helper cells/cell mediated immunity
(TH) or cytotoxic (TC)

• Platelets – 15,000 to 400,000; involved in blood clotting, are not actually cells,
are fragments of cells

Phagocytosis – means in Greek: eat & cell; ingestion of microorganism or cellular debris
by microorganism

Composition of blood:
Plasma – blood consists of a fluid called “this”, which contains formed elements (cells
and cell fragments)

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Cells – Red Blood Cells (RBC), White Blood Cells (WBC), Platelets

Leukocytosis – increase in WBC count that occurs during:

• mononucleosis
• appendicitis
• pheumonia

Leukopenia – drop in WBC count that occurs during:

• HIV/AIDS infection

Differential Blood Count – a blood test showing percentage of each kind of white blood
cell in a sample of 100 WBCs

Leukocytes (WBC):
• Granulocytes:
o Neutrophils – stains with acid/basic dyes; also called PMNS
(PolyMorphoNuclearLeukocytes)

Highly Motile

Phagocytic in initial stages of infection
o Basophils – stains with basic dyes; produces histamine involved in
inflammation and allergic reactions
o Eosinophils – stain with an acid dye called eosin; are somewhat
phagocytic; produce toxic substances against large parasites like
helminthes

Also produce peroxide ions after attaching to the outside surface of
parasite
• Agranulorcytes:
o Monocytes – when activated mature to macrophages and their function is
phagocytosis

Also take part in Specific Cell Mediate Immunity
o Lymphocytes – present in blood and lymphatic system (Tonsils, Spleen,
Thymus, Peyers Patches, Red Bone Marrow)

B Cells

T Cells

Lymphatic System – the fluid flowing between tissue cells is picked up by lymphatic
vessels and brought into “this”

Actions of Phagocytic Cells:

• Both of these mature to macrophages
o migrating monosites
o fixed macrophages

• During infection, neutrophils initially dominate, and later on macrophages

dominate

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Mechanism:
1) Chemotaxis – is the chemical attraction of phagocytes to microorganisms, white
blood cell secretions; compliment activation
2) Opsonization – first the foreign microorganisms are coated with serum proteins,
this process is called “this”, so that phagocytes can better adhere/attach to the
microorganism; compliment proteins take part in this activity and are called:
a. opsonins
3) Ingestion – pseudopod formation which engulfs the microorganism, a phagosome
forms and is highly acidic and digests the pathogen
4) Digestion – phagosome (vacuole forming around the pathogen) contacts the
lysosome organelle aiding in “this”
a. lysomes – contain digestive enzymes and bactericidal substances
5) Exocytosis – ejection of residues

Other Important Chemicals:

• nitric-oxide – involved in inflammation

Inflammation – is a defensive mechanism triggered by damage; causes:

• redness
• pain
• heat
• swelling
• sometimes loss of function

Acute Inflammation – very intense and short-lived; proteins are produced by the liver,
which include:
• complement proteins
• cytokines – language of cells
• kinins – helps in for vasodilation
• fibrinogen – helps in for clotting

Chronic Inflammation – less intense and long-lasting and is more damaging, can lead to
arthritis and other diseases

Function of Inflammation:
• destroys injurious agent
• confines infection
• repair

vasodilation – increased permeability of blood vessels thus increasing blood flow to the
damaged area
• results in redness (erythema) and heat
• edema – swelling of inflamed region

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During Tissue Damage:
• blood vessels dilate (vasodilation)
• their permeability increases

Histamines – are produced by Basophiles; cause vasodilation

Kinin – causes vasodilation and increase permeability and chemotaxis

Prostaglandins – substances released by damaged cells, which intensify the effects of

histamines and kinins; help phagocytes to move through walls of blood vessels

Leukotrienes – are substances produced by mast cells and basophils; increase

permeability of blood vessels and Chemotaxis and Opsonization

After/Within One Hour of Injury/Inflammation; Phagocytes appear:

1) margination – Neutrophils and Monocytes stick to capillary walls
2) diapedesis - phagocytes squeeze between endothelial cells to reach the damaged
area; emigration occurring within 2 minutes
3) phagocytosis – phagocytes begin to destroy invading microorganisms by “this”
process

• inflammation is local
• fever is systemic

hypothalamus – fever is controlled by the “this”

Lipopolysaccharides (LPS)

Interleukin-1 (IL-1) – interacts with prostaglandins causing the hypothalamus to reset

body temperature

Fever:
• increases metabolism
• blood vessels contract
• shivering
• up to a point, fever is considered bodily defense
• It intensifies action of Interferons and production of T-Lymphocytes

Interferons – an antiviral protein produced by certain animal cells in response to a viral

infection; are chemicals produced by virally infected cells which goes to uninfected cells
which results in those cells producing chemicals which prevent viral infection to them

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Complement System – is a defensive system consisting of over 30 proteins produced by
the liver and found circulating in blood serum and within tissues throughout the body
• its function is to destroy microbes by:
o cytolysis
o inflammation
o phagocytosis induction

• complement proteins are designated by uppercase “C” and are inactive till it is
cleaved into products
• proteins are numbered C1 – C9 p.473
• active proteins are indicated by lowercase a & b
o for example, inactive complement protein C3 is cleaved resulting in active
proteins:

C3a

C3b

Complement Functions:
• opsomization – coating microbes with proteins, resulting in phagocytosis
• cytolysis – membrane attack complex (MAC)  pores in cells  resulting in
“this”; which is the destruction of cells, resulting from damage to their cell
membrane, that causes cellular contents to leak out
• Membrane Attack Complex – complement proteins C5 – C9, which together
make lesions in cell membranes that lead to cell death
• histamines (Mast Cells)  inflammation

Compliment Activation:
• C3 is the most important because it starts a cascade that results in cytolysis,
inflammation and phagocytosis
• C3 is split into activated compliment proteins:
• C3a – acts in inflammation
• C3b – enhances phagocytosis
i. Cascade – complement proteins act in a “this”, that is, one
reactions triggers another, which, in turn, triggers another, and so
on
• C3b connects with C5 and opsomization occurs
• C6, C7, C8, C9  Cytolysis

Three Complement Pathways:

I. Classical – Antibody Mediated:
• C1 cleaves interacting with C2 and C4, which interact with C3 resulting in
opsomization, cytolysis (MAC) and Inflammation
II. Alternative – B, D, P Protein Mediated:
• B, D, P interact with C3 protein restuling in opsomization, cytolysis
(MAC) and inflammation
III. Lectin/Mannose Mediated:

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 • Lectin/Mannose interact with C2 and C4, which interact with C3 resulting
in opsomization, cytolysis (MAC) and Inflammation

Chapter 17: Specific Defenses of the Hosts: The Immune

Response
Acquired (Adaptive) Immunity:

Antigen – any substance which provokes a specific response

• example: antibody production, activation of lymphocytes and other cells
like macrophages

Naturally Acquired Immunity:

• active – bodily production of antibodies
• passive – mother produces antibodies which are passed on to a fetus

Artificially Acquire Immunity:

• active – vaccination
• passive – during blood transfusion; antibody is in the blood serum

antiserum – blood-cell derived fluid with antibodies

serology – study of antigens, antibodies and their subsequent reactions

immunoglobulins – antibodies are also called “this”

Immunoglobulins consist of three proteins:

1) Alpha
2) Beta
3) Gamma

Structure of an Immunoglobulin:
• two heavy chains
• two light chains
• disulfide linkage
• is a Y shaped molecule which can turn into a T shaped molecule

Specific Immunity:
• Humoral – immunity is fluid based; consists of antibodies in the serum
o B-Cells – these antibodies are produced by “these”;

Plasma Cells – B-cells mature to form “this”; which
produce antibodies

Stem Cells – B-Cells are developed from “this” within red-
bone marrow

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T-Cells – B-Cell stimulation is assisted by “this”

Apoptosis – programmed cell death, if B-Cells do not
encounter antigens, will experience “this”
• Caspaces – enzymes that destroys a cell by
destroying the proteins within the cell
• Cell-Mediated – immunity is cell-based; consists of antibodies in the cell
o T-Cells:

Macrophages
• Monocytes – “these” mature to form macrphages

B-Cells

Antigens – are made of carbohydrates and proteins from viruses

Epitopes – are highly specific regions on antigens which are antigenic determinants;
meaning they are recognized by antibodies

• antibodies connect to antigens

Hapten – small antigens; are less than 10,000 daltons; attached to very large proteins to
initiate an immune response

Ig – stands for Immunoglobulin

Five Different Ig Classes:

1) IgG (most common)
2) IgM
IgA
3) IgD
4) IgE

AB – stands for Antibody

• about 10 million cells are made each day

Activation of Antibody Producing Cells by Clonal Selection:

• each B-Cell carries a specific antigen recognizing receptor
• clone selection – once specific antigen binds, than that leads to “this”
• clone amplification – one clone selection occurs, the spread or increase of
B-Cells occur called “this”
• Mature B-Cells
o Memory B-Cells – can last for 10 years
o Plasma Cells – produces antibodies

After Antigen/Antibody formation:

compliment proteins - phagocytosis is facilitated by “this”
• opsenization

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 • cytolysis – MAC causes “this”
• inflammation

Polyclonal Antibodies – different B-Cells producing antibodies against the same protein
and may recognize the same or different epitopes
• antibodies are from multiple clones of B-Cells

Monoclonal Antibodies – are derived from a single B-Cell and very highly specific
recognizing a single epitope

Monoclonal Antibody Production Steps:

1) injection of antigen into a mouse
2) sacrifice the infected mouse
3) procure spleen B-cells
4) serial dilution
5) One cell is separated
6) Spleen B-Cell is fused with a Tumor Cell , which is immortal
7) A new cell is produced producing antibodies, which is immortal since it was
derived from a tumor cell

• Humoral Immunity is not a vigorous immune response

• Cell Mediated Immunity is more vigorous than Humoral

• Cell Mediated Response is the chief immune response

Language of Immune Cells:

Cytokines – can cause:

• B-Cell activation
• B-Cell differentiation – B-Cells changing to plasma cells

Interleukins (IL) – cytokines produced by leukocytes are called “this”

• there are at least 26 interleukins

IL-1 – is produced by every nucleated cell (stimulates T-Cells)

IL-2 – T-Cells (activation of B-Cells, NK Cells, Tc Activiation

Different types of Cytokines:

• TNF – important in inflammatory response
• CSF – development of blood cells
• GMCSF – stimulates formation of red blood cells and white blood cells

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Cellular Components of Immunity:
• T-Cells develop from stem cells

Thymus – T-Cells develop in bone marrow and mature in “this”

Pathogen:
1) T-Cell Receptor
a. Memory cells
b. Activated T-Cells (by clonal selection)
2) Direct Recognition of Pathogen by dendritic cells or macrophages (by APC –
Antigen Presenting Cells)

Four Types of T-Cells:

1) TH – T-Helper
2) TC – T-Cytotoxic
3) TD – Delayed Hypersensitivity
4) TS – Suppressed T-Cells

• TD and TS are mixtures of TH and TC

Cluster of Differentiation (CD) (receptors on T-Cells):

CD4 – TH are also called “this”

CD8 – TC are also called “this”

Antigen Presenting Cells (APG) – Ingests/Digests pathogen so that the pathogen is now
a 10 amino acid peptide, which is the target
• by itself, the digested peptide will not initiate an immune response
• therefore, it needs to be introduced by special proteins called Major
Histocompatibility/Protein Complex (MHC)

MHC – introduces the 10 amino acid peptide to immune system

Steps to Cell Mediated Immunity:

1) pathogen is ingested by APC (Antigen Presenting Cell)
2) pathogen is digested to a 10 amino acid peptide
3) MHC pushes 10 amino acid peptide out of the APC, and introduces the 10 amino
acid peptide to TH Cells
4) IL-1 is produced by APC,
5) TH Cells produces IL-2

• TH is the Heart of Immunity

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Types of TH Cells:
• TH1 – produces cytokines that activate cells related to cell-mediated immunity
(macrophages, CD8, NK Cells)
• TH2 – activates B-Cells

Non-Specific Cellular Components:

1) activated macrophages
2) Natural Killer (NK) Cells

Antibody Dependent Cell-Mediated Cytotoxicity (ADCC) – antibody and cytotoxic

cells are used in a response specific for very large pathogens like a Helminthes

Steps:
1) coating of a large pathogen with antibody
2) NK Cells (produce perforin), macrophages, neutrophils, eosinophils all bind to the
Antibody Fc
3) Lysis by substances produced (peroxide and perforins)

The duality of the immune system: p.503

1) Humoral Immunity
2) Cell Mediated Immunity

Chapter 19: Disorders Associated with the Immune System

AIDS:
• 1981 – Pneumonia cases in LA, these people were severely
immunocompromised
o These patents had cancer of blood and skin cells – Kaposi sarcomas

Montagnier and Robert Gallo – discovered HIV 1

Origin of HIV:
• viruses which infected chimps and monkeys in Africa and eventually they
crossed species into humans

Structure of HIV:
• is an Envelope Virus
• the envelope has several spikes called GP120, which are proteins for viral entry
• has 9 genes expressed by the RNA Genome
• reverse transcriptase

gp120 and gp41 – essential proteins allowing viruses to penetrate a host cell

GAG – viral envelope and capsule

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POL – RT, integrase, protease

Other Genes:
• TAT – very important for viral replication
• NEF – regulatory protein
• REV – regulatory protein
• VPR – regulatory protein
• VPU – particle release
• VIF – viral infectivity

HIV Life Cycle:

• a very important part is viral entry
• HIV – 1 primary infects Helper T-Cells (TH)
• CD4 receptor – HIV Virus binds to the receptor of T-Cells which are called the
“this”
• CD4 receptors interact with gp120 and gp41 proteins
• CCR5 and CXCR4 also interact with gp120 and gp41 proteins
• Once virus enters cell, reverse transcribes RNA to form DNA
• Provirus  integrate in genome, production of viral particles

Provirus – HIV integrated DNA may not produce new HIV but remains hidden in the
host cell’s chromosome as a “this”

Virus is quite elusive where it:

1) hides as provirus
2) virus then can remain in memory T-Cells and hide there
3) they can fuse with other cells and replicate without being released
4) phenomenal mutation rate
5) targets heart of immunity

Prevention:
• safe sexual practices
• treatment – AZT, HIV Vaccinations

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 Microbiology Test 2 Questions
1) How do bacteria protect their genomic DNA from restriction enzymes?

Methylates – addition of a methyl group (-CH3) to a molecule;

o bacterial DNA can be protected from digestion/enzyme cutting because

the cell methylates some of the cytosines in its DNA

2) cDNA Library is more useful than genomic DNA Library; Why?

o Cause all the introns are taken out.

o from this process, genes lacking introns can be isolated and made
o this library is used to make proteins

3) Why are shuttle vectors useful?

shuttle vectors – a plasmid that can exist in several different species; used in
genetic engineering

4) Bacterial Transformation – the process in which genes are transferred from one
bacterium to another as “naked” DNA in solution; DNA is precipitated with
calcium phosphate and applied to cells (used for bacterial and eukaryotic cells)
o some lipid based chemicals also used

5) Given a protein sequence, can you deduce the DNA sequence?

No, due to degeneration of code

6) Southern Blotting – a technique for fingerprinting that uses DNA probes to

detect the presence of specific DNA in restriction fragments separated by
electrophoresis

7) Gives you diagrams; Restriction Enzyme Analysis – Cutting a plasmid

strand and analyzing a molecular weight chart.

Restriction enzyme – an enzyme that cuts double-stranded DNA at specific sites

between nucleotides

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 o Recognition sites (such as GAATTC) have to be copied and equal
o a restriction enzyme recognizes and cuts, or digests, only one particular
sequence of nucleotide bases in DNA, and it cuts this sequence in the
same way each time
o restriction enzymes used in cloning experiments recognize four-, six- or
eight-base sequences

8) Gives you diagrams; Restriction Pattern (Same kind of question like 7)

Example: An enzyme that cuts double stranded DNA at specific sites between
nucleotides 30 = 5, 2, 7 14

9) Is about PCR (Polymerase Chain Reaction) Is a polymerase chain reaction

possible in a bacterial polymerase chain?

No, because high temperature will kill the bacteria

Polymerase Chain Reaction (PCR) – is a technique by which small samples of

DNA can be quickly amplified, that is, increased to quantities that are large
enough for analysis

Steps:
• 1) incubate target DNA at 94 degrees Celsius for 1 minute to separate
strands (denaturation)
• 2) add primers, nucleotides (deoxynucleotides A, C, G, T) and DNA
polymerase
• 3) Primers attach to single-stranded DNA during incubation at 60 degrees
Celsius for 1 minute
• 4) incubate at 72 degrees Celsius for one minute; during this time, two
copies of target DNA are formed
• 5) repeat the cycle of heating and cooling to make two more copies of
target DNA

70 degrees Celsius polymerase activity

annealing temperature (between45 degree and 70 degree Celsius
variable)

10) Is also about PCR

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 11) How is a phage library constructed?

o the goal is to make a collection of clones large enough to ensure that at

least one clone exists for every gene in the organism

ICVT – international committee on taxonomy of viruses

ICVT groups viruses into families based on:

• nucleic acid type
• strategy for replication
• morphology

virus – genus names end in “this”

viridae – family names end in “this”
ales – order names end in “this”

12) Two techniques of Clone Selection: Colony Hybridization and Beta-Gal

Selection

Two Methods for selecting a clone:

Blue-White Selection – using β-Galactosidase with X-Gal, bacteria turns
blue; however when the β-Galactosidase is disrupted , the bacteria turns white,
disruption caused by the fact that a gene was inserted into the β-Galactosidase
i. Plasmid DNA and foreign DNA are both cut with the same
restriction enzyme

a. Foreign DNA is inserted into the plasmid, where it inactivates the lacZ
gene
b. The recombinant plasmid is introduced into a bacterium, which
becomes ampicillin-resistant
c. All treated bacteria are spread on a nutrient agar plate containing
ampicillin and β-Galactosidase substrate, and incubated
d. White colonies that appear must contain foreign DNA
e. Blue colonies must not contain foreign DNA
• It is not a sensitive method and there can be background

Colony Hybridization – the identification of a colony containing a desired gene

by using a DNA probe that is complementary to that gene; highly selective
method which is highly sensitive
a. Create a master plate with colonies of bacteria containing cloned
segments of foreign genes
b. Make replica of master plate on nitrocellulose filter paper by placing the
filter paper on top of the master plate

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 c. Treat filter paper with detergent (SDS) to lyse bacteria
d. Treat filter paper with sodium hydroxide (NaOH) to separate DNA into
single strands
e. Add radioactively labeled probes
f. Probe will hybridize with desired gene from bacterial cells
g. Wash filter paper to remove unbound probe and expose filter paper to x-
Ray film
h. Developed film is compared with replica of master plate to identify
colonies containing gene of interest

13) How are recombinant proteins produced?

Steps for Recombinant DNA Procedure:

1. Vector such as a plasmid is isolated
2. Plasmid – a small circular DNA molecule that replicates independently of
the chromosome
3. DNA is cleaved by an enzyme into fragments
4. gene fragment of interest is inserted into plasmid
5. plasmid is taken up by a cell such as a bacterium; plasmid multiplies
6. cells with gene of interest are cultured and then cloned
i. Clone – a population of cells arising from a single parent cell
7. Two possibilities:
i. copies of gene are harvested
ii. bacterial cells make a protein product
o copies of protein are harvested

cDNA libraries – contain copies of genes made from mRNA

a. circumvent introns
b. take mRNA (lacks introns) through isolation, invoke reverse transcription
resulting in complementary DNA (cDNA); than make a double-stranded
cDNA and than clone into bacteriophage vectors
c. from this process, genes lacking introns can be isolated and made
d. this library is used to make proteins

14) How are subunit vaccines produced?

subunit vaccines – consisting only of a protein portion of a pathogen, are being

made by genetically engineering yeasts

15) How were restriction enzymes discovered?

• they were discovered when certain bacteriophages were found to have

a restricted host range

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 • if these phages were used to infect bacteria other than their usual hosts,
restriction enzymes in the new host destroyed almost all the phage
DNA

16) Recognize the complimentary sequence?

For DNA:
• Adenine bonds with Thymine
• Cytosine bonds with Guanine

For example: GGCTACCC…so compliment is CCGATGGG

17) electroporation – a technique by which DNA is inserted into a cell using an

electric current; electrocuting cells in the presence of DNA

18) What are the uses of synthetic DNA?

Synthetic DNA – under certain circumstances, “this” can be made in vitro with
the help of DNA synthesis machines; outside of the cell chemically

o can be used to make entire genes like insulin

19) Know technique of Colony Hybridization – clone selecting techniques

Colony Hybridization – the identification of a colony containing a desired gene

by using a DNA probe that is complementary to that gene; highly selective
method which is highly sensitive
a. ate a master plate with colonies of bacteria containing cloned segments of
foreign genes
b. Make replica of master plate on nitrocellulose filter paper by placing the
filter paper on top of the master plate
c. Treat filter paper with detergent (SDS) to lyse bacteria
d. Treat filter paper with sodium hydroxide (NaOH) to separate DNA into
single strands
e. Add radioactively labeled probes
f. Probe will hybridize with desired gene from bacterial cells
g. Wash filter paper to remove unbound probe and expose filter paper to x-
Ray film
h. Developed film is compared with replica of master plate to identify
colonies containing gene of interest

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 20) Which is the most frequently mutated gene in human cancer is the?

P53 Gene

21) Human Genome Project, what is it?

Human Genome Project – the goal of this project is to map the 35,000-70,000
genes in human DNA and to sequence the entire genome, approximately 3 billion
nucleotide pairs
o was started to map and sequence the entire human genome

sequence – to formulate the exact order of nucleotides

22) What is not true about the Human Genome? (so know what is true of the
human genome)
a. Has 3 billion bases
b. 90% noncoding, 10% coding
c. Condense into chromosomes
d. There are 35,000 – 70,000 genes
e. Francis Collins – Human Genome Project director

23) Restriction Fragment Length Polymorphism (RFLP) – a fragment resulting

from restriction-enzyme digestion of DNA during the Southern Blotting technique

24) anti-sense technique – is a technique for inhibiting gene expression at the RNA
level

25) Plasmids and Transposons: how do they differ?

• Plasmid – a small circular DNA molecule that replicates independently of

the chromosome
• Transposon – a small piece of DNA that can move from one DNA
molecule to another

26) Daughter cells are most likely to inherent which one of the following from the
parent cell?

o Any change in DNA is inherited, not RNA

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 27) Gives you a diagram, plasmid with some cloning site and a fragment of DNA,
can it be inserted and cloned?

o Polar side enzymes of the fragment must match up with the polar ends of
the fragment enzyme area to insert

28) DNA sequencing – the process by which the nucleotide sequence of DNA is
determined

29) What does Virus nucleic acid consist of?

o Contain a single type of nucleic acid, either RNA or DNA as genome

30) Viral Protein Coating is called?

o Capsid – the protein coat of a virus that surrounds the nucleic acid

capsomere – a protein subunit of a viral capsid

31) The host of a phage are?

o Bacteria

32) What is Lysogenic and Lytic Lifecycles?

Life Cycle of a Phage:

Two Different Life Cycles:
• Lytic Cycle – ends with the bacteria lysing and the viruses escaping from the
lysed bacteria
a. Steps:
i. Attachment – phage attaches to host cell
ii. Penetration – phage penetrates host cell and injects its DNA
iii. Biosynthesis – Phage DNA directs synthesis of viral components
by the host cell
iv. Maturation – viral components are assembled into virions
v. Release – host cell lyses and new virions are released

• Lysogenic Cycle – bacteria remains alive, the virus forms a symbiotic

relationship with the bacteria
a. Steps:
i. Attachment – phage attaches to host cell
ii. Penetration – phage penetrates host cell and injects its DNA

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 iii. Viral DNA integration into bacterial genome
iv. Infected bacteria goes on replicating more infected bacteria

33) Virus infecting a bacterial cell creates a pore by?

• By an enzyme secretion:
o Lysozyme – an enzyme capable of hydrolyzing bacterial cell walls

34) Know something about HIV-1/Reverse Transcriptase: Does it have proof

reading activity?

o No, because HIV mutates at a high rate

35) Orthopoxvirus:

o Viral Family – Poxviridae

o Characteristic – Double-stranded DNA enveloped 200 – 350 nm
o Clinical Features – causes diseases such as smallpox, malluscum
contagiosum and cowpox

36) Rhinovirus:

o Viral Family – Picornaviridae

o Characteristic – Single-stranded RNA enveloped 28-30nm
o Clinical Features – more than 100 Rhinoviruses exist and are the most
common cause of colds

37) Heptitis B

o Viral Family – Hepadnaviridae

o Characteristic – 42 nm
o Clinical Features – after protein synthesis, hepatitis B viruses uses
reverse transcriptase to produced DNA from mRNA; causes hepatitis
B and liver tumors

38) Prions propagate/multiplies?

• Prions – proteinacesus infectious particle

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 • Results in conversion a normal host glycoprotein (PrPc) for cellular prion
protein into an infectious form called PrPsc for scrapie-protein causing Mad
Cow Disease

Steps to Mad Cow Disease infection:

1. PrPC is located on chromosome 20 in humans and is secreted to the cell
surface
2. PrPSc reacts with PrPC on the cell surface
3. PrPC converts to PrPSc
4. PrPSc is taken in by endocytosis and accumulates in lysosomes resulting
in the eventual formation of Mad Cow Disease

39) HIV Virion contains? and what is the structure?

Structure of HIV:
o is an Envelope Virus
o the envelope has several spikes called GP120, which are proteins for viral
entry
o has 9 genes expressed by the RNA Genome
o reverse transcriptase

gp120 and gp41 – essential proteins allowing viruses to penetrate a host cell

40) HIV gains entry in cells by?

• CCR5, CXCR4, GP120, GP41

41) HIV can evade detection for decades by?

• Provirus – HIV produced by a host cell is not necessarily released from the
cell but may remain as latent virions in vacuoles within the cell

42) HIV is difficult to eradicate; why?

Virus is quite elusive where it:

o hides as provirus
o virus then can remain in memory T-Cells and hide there
o they can fuse with other cells and replicate without being released
o phenomenal mutation rate
o targets heart of immunity: T-Cells and Macrophages

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 43) HIV-TAT protein; what is the use of it?

• viral replication

44) What is the function of HIV protease?

o HIV Protease - The active site of the HIV protease binds to the
polyproteins and cleaves them into functional proteins essential to the
structure of HIV

o HIV must use a HIV encoded enzyme called protease in order to cleave a
large Gag-Pol polyprotein (p120), a Gag polyprotein (p55), and an Env
polyprotein (gp160) into functional proteins essential to the structure of
HIV and to its RNA packaging. The active site of the HIV protease binds
to the polyproteins and cleaves them into functional proteins

45) What of the functions in immunity is not a mechanical factor?

Mechanical factors are:

o skin
o mucous membranes

mucous
o Lacrimal apparatus – protects the eyes by producing tears and
pathogenic microbes are washed away or digested by an enzyme called
lysozyme
o saliva

46) What of the following has the highest phagocytic activity?

o Neutrophils

47) Which is not inflammation? (know what is inflammation)

Inflammation – is a defensive mechanism triggered by damage; causes:

o redness
o pain
o heat
o swelling
o sometimes loss of function

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 Function of Inflammation:
o destroys injurious agent
o confines infection
o repair

vasodilation – increased permeability of blood vessels thus increasing blood flow

to the damaged area
o results in redness (erythema) and heat
o edema – swelling of inflamed region

During Tissue Damage:

o blood vessels dilate (vasodilation)
o their permeability increases

Histamines – are produced by Basophiles; cause vasodilation

Kinin – causes vasodilation and increase permeability and chemotaxis

48) Differential Blood Count – a blood test showing percentage of each kind of
white blood cell in a sample of 100 WBCs

o refers to white blood cell count explicitly, calculation of the present age of
each kind of white cell in a sample of 100 white blood cells

49) What causes vasodilation?

o Histamines

50) Which of the following is true of an interferon; what is an interferon?

Interferons – a natural glycoprotein formed by cells exposed to a virus,

introduces production of translation inhibitory protein (TIP), which blocks
translation of viral ribonucleic acid giving other cells protection

o an antiviral protein produced by certain animal cells in response to a viral

infection; are chemicals produced by virally infected cells which goes to
uninfected cells which results

51) What is most found in the serum?

o Complement proteins

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 52) What is an effect of opsonization?

Opsonization – first the foreign microorganisms are coated with serum proteins,
this process is called “this”, so that phagocytes can better adhere/attach to the
microorganism; compliment proteins take part in this activity and are called:
o opsonins

53) Classical pathway, how is it initiated?

Classical – Antibody Mediated:

• Activated C1 cleaves interacting with C2 and C4 by splitting them, which
interact with C3 resulting in opsomization, cytolysis (MAC) and
Inflammation

54) What is a function of membrane attack complex (MAC)?

Complement System – is a defensive system consisting of over 30 proteins

produced by the liver and found circulating in blood serum and within tissues
throughout the body
o its function is to destroy microbes by:

cytolysis

inflammation

phagocytosis induction

• Membrane Attack Complex – complement proteins C5 – C9, which

together make lesions in cell membranes that lead to cell death

• cytolysis – membrane attack complex (MAC)  causes pores in cells 

resulting in “this”; which is the destruction of cells, resulting from damage
to their cell membrane, that causes cellular contents to leak out

55) Which of the following is involved in specific immunity?

• lymphocytes

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 56) Which of the following is involved in the resistance to parasite helminthes?

Antibody Dependent Cell-Mediated Cytotoxicity (ADCC) – antibody and

cytotoxic cells are used in a response specific for very large pathogens like a
Helminthes
Steps:
1) coating of a large pathogen with antibody
2) NK Cells (produce perforin), macrophages, neutrophils,
eosinophils all bind to the Antibody Fc
3) Lysis by substances produced (peroxide and perforins)

57) Macrophages arise from which of the following?

• Monocytes – “these” mature to form macrphages

58) In addition to lymphocytes, which cells do not have visible granules in their
cytoplasms?

• Monocytes

59) Which is the most important compliment protein?

• C3

60) Which of the following is not true about the classical pathway of compliment
activation?

Classic Pathway Steps:

• a pair of antibody molecules attaches to an antigen, for example, proteins
or larger polysaccharides on the surface of a bacterium or other cell,
forming an antigen-antibody complex
• the antigen-antibody complex binds and activates C1
• C1 activates C4 and C2 by splitting them. C4 is split into fragments called
C4a and C4b, and C2 into fragments called C2a and C2b
• C4b and C2b combine and together they activate C3 by splitting it into
C3a and C3b. The C3 fragments then initiate cytolysis, inflammation and
opsonization

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 61) The specificity of an antibody lies in which region?

• Antigenic Determinants (epitopes) – a specific region on the surface of an

antigen against which antibodies are formed
• the nature of this interaction depends on the size, shape and chemical
nature of the antigenic determinant, as well as the chemical structure of
the binding site on the antibody molecule

62) B-Cells mature and become?

• Plasma cells

63) Antigen is?

• Antigen – a chemical substance that causes the body to produce specific
antibodies or sensitized T-Cells
• Anything that promotes a response from the immune system
• it is not necessarily a foreign body

64) Which of the follow destroys virus infected cells?

• Cytotoxic T Cells – destroy target cells on contact; a specialized T Cell

that destroys infected cells presenting antigens
• TC

65) Which of the follow is involved in antibody production?

• Helper T Cells – immune system cells that play a central roll in the
immune response; a specialized T Cell that often interacts with an antigen
before B Cells interact with the antigen
• TH

66) Which chemical causes transmembrane channels in target cells?

• perforins

67) All of the follow is true about Natural Killers (NK Cells) except?

Natural Killer (NK) Cell – a lymphoid cell that destroys tumor cells and virus-
infected cells
• they are not T-Cells and are not antigenically specific

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 68) Monoclonal antibodies against CD4 antigens might be used to treat AIDS
because?

Monoclonal Antibodies – are derived from a single B-Cell and very highly
specific recognizing a single epitope

Monoclonal Antibody Production Steps:

1. injection of antigen into a mouse
2. sacrifice the infected mouse
3. procure spleen B-cells
4. serial dilution
5. One cell is separated
6. Spleen B-Cell is fused with a Tumor Cell , which is immortal
7. A new cell is produced producing antibodies, which is immortal since it
was derived from a tumor cell

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