IMMUNOTOXICOLOGY

BAG / SMF. Ilmu Kesehatan Kulit &

Kelamin
FK.UNS/ RSUD.dr.Moe3wardi
SOLO

Immunotoxicology
Study of the effects of physical and
chemical agents on the immune system
Immune system is highly sensitive to
toxicants

Consists of many cells undergoing rapid

proliferation and differentiation;
(eg. Bone marrow)
Immunocompetence requires adequate
balance (cooperation) and functioning
between different cellular components

Functions of the Immune System

Defense

Homeostasis

Surveillance

Anatomical Organization

External Secretory Systems

Respiratory, GI, urogenital

Internal Secretory Systems

Lymph system
Primary lymphatic organs (thymus, fetal liver,
bone marrow)
Secondary lymphatic organs (spleen, lymph
nodes, peritoneal cavity, lymph ducts)
Reticuloendothelial system (mononuclear
phagocytes; macrophages, monocytes)

INNATE IMMUNITY

PHYSICAL
BARRIERS
Skin, mucous
membrane

CELLS
granulocytes,
monocytes,
macrophages

CHEMICAL
BARRIERS
pH, lipids, enzymes

ADAPTIVE IMMUNITY
HUMORAL
B cells
antibodies

CELL MEDIATED

MP

T cells
lymphokines

Cellular Components of the

Immune System

Neutrophils and Macrophages

Nonspecific (innate) immunity

Secrete biological mediators
Process and present antigen (MP)
Pattern recognition receptors; Fc receptors for
antibody coated antigens (MP)

Lymphocytes

Specific (acquired) immunity

Specific antigen receptors
B cells (humoral immunity)
T cells (cell mediated immunity)

Cellular Interactions in the

Immune System
Activated
MP

MP
PA

MP
AG

(10%)

(+)

(+)

(+)
Sensitized

Th

Th

(-)

Antigen
destruction

(+)

Tcyt

(90%)

Treg

(-)
Sensitized

(-)
plasma
Bmem

Hematopoietic Bone Marrow Stem Cells

Stem cells
can differentiate
into many
different cell types
How does it
decide what
to become?

Control of Stem Cell Differentiation

Two major influences:

1. ENVIRONMENT in which it develops: which
primary lymphoid organ (ie bone marrow vs
thymus)
2. CYTOKINES AND GROWTH FACTORS stem
cells are exposed to during this process

Origin and Development of

Immune Cells
BONE MARROW
Stem
cells

noncommitted
dividing

Lymphoid

Progenitor
cells
committed
dividing

BLOOD
T cell

thymus

90%

TISSUE
Cell Mediated
Immunity

30%

bursa
equivalent

B cell
10%

Humoral
Immunity

Monocyte
3-5%

Myeloid

Granulocyte

MP

PMN (70%)
eosinophil (<3%)
basophil (<2%)

mast

Innate Immunity

Universal and evolutionarily conserved mechanism

of host defense against infection; first line of
defense;
Predates adaptive immune response
Found in all multicellular organisms
(adaptive only in vertebrates)
Uses receptors and effectors that are ancient in
their lineage
Provides protection against a wide variety of
pathogens (pattern recognition receptors)
Distinguishes self from non-self perfectly
Defects in innate immunity are very rare and almost
always lethal

Elie Metchnikoff: Father of Macrophages

1908 Nobel prize
Metchnikoff watched the reaction to a splinter inserted into a
starfish. Hemocytes (amoeba-like cells) arrived and tried to
ingest the foreign body; if they could not, they walled it off.
Mammals do exactly the same thing to foreign bodies.

Phagocytosis- to devour

Oyster hemocyte

Mouse macrophage

The similarity between an invertebrate and mammalian phagocyte is

striking; they also use many of the same mechanisms, including the
production of reactive oxygen species
http://www.mdsg.umd.edu/oysters/oysblood.htm

http://itgmv1.fzk.de/www/itg/diabate/images.html#fig5

Foreign Body Reactions in Humans

chromic catgut

The suture material is not digestible by

macrophages (MP), so it has been walled
off by fibroblasts.
Atlas of Granulomatous Diseases Yale Rosen, M.D.

silicone droplets

The clear (unstained) globules of silicone have

been released from a ruptured breast implant.
They cannot be ingested; at this early stage,
MP accumulate to wall off the material; note MP
that has become a multinucleated giant cell
showing the asteroid bodies characteristic of
the foreign body reaction.

Innate Immunity: Functions

Provides a barrier to prevent the

spread of infection

Mechanical (tight junctions, movement)

Chemical (fatty acids, enzymes, pH,
antimicrobial peptides)
Microbiological (normal flora)
Mucosal surfaces

Respiratory, GI, Reproductive

Skin (epithelial cells)

Wounds, burns, insect bites

Innate Immunity: Functions

Identifies and eliminates pathogens

Non-adaptive recognition systems
Activates molecules that target the
microbe and aid in its identification
These factors may be surface
expressed, released from immune
cells or present within circulatory
system

Innate Immunity: Functions

Initiates an inflammatory response

Reaction to injury or infection
Trauma to tissues or cells
Presence of foreign material (non-self)
Infectious agents (viruses, bacteria, fungi)

Delivers effector molecules and immune

cells to the site of injury/infection
Components
Granulocytes, MP, secreted factors
Blood vessels (endothelium)
Plasma proteins

Innate Immunity: Functions

Provides signals to alert the

adaptive immune system to
activate an effective specific
immune response
Upregulation of co-stimulatory
molecules MHC class II, CD80/86
Induction of cytokine/chemokine
response : IL-4, IL-12

Innate Immunity:
Cellular Components

Granulocytes
Polymorphonuclear leukocytes
(PMN, neutrophils)

Eosinophils
Basophils (blood)
Mast Cells (tissues)

fight
pathogens
inflammation

allergic and
hypersensitivity reactions

Mononuclear Phagocytes (RES)

Monocytes (blood)
Macrophages (tissue)

fight pathogens
inflammation
cytotoxicity
immune regulation

Innate Immune Cells

Origin and Development of

Macrophages and Granulocytes
Bone Marrow
monoblast

Blood
promonocyte

Tissue

monocyte
macrophage

Phagocyti
c
precursor
s

basophilic
promyelocyte

basophil

eosinophilic
promyelocyte

eosinophil

neutrophilic
promyelocyte

Neutrophil
(PMN)

myeloblast

mast
cell

Neutrophils
(PMN)

Present in blood (60-70% of WBC)

Not normally present in tissues
Short lifespan - 12 hours
Functions:
First cell at the site of infection/injury

Ingest and kill microbes after bactericidal

mechanisms activated (binding to pathogen)

Mononuclear
Phagocytes

Blood - monocytes (1-6% WBC)

Tissues - macrophages
mature form of monocytes
found in tissues (ex., gastrointestinal tract, lung, liver,
brain, skin, spleen); reticuloendothelial system (RES)

Functions:
Phagocytize and kill after bactericidal mechanisms
activated
Produce cytokines/chemokines (initiate inflammation)
Antigen presentation (activate adaptive immunity)
Tumor surveillance and cytotoxicity

Reticuloendothelial System

Consists of fixed and wandering

macrophages located throughout the body
(tissues, sinusoids, lymph system)
Major locations: Liver and Lung
Functions: antibacterial resistance, tumor
resistance, defense against shock, antigen
processing, lipid metabolism, protein
turnover, iron metabolism

Origin and Development of

Macrophages and Granulocytes
Bone Marrow
monoblast

Blood
promonocyte

Tissue

monocyte
macrophage

Phagocytic
precursors

basophilic
promyelocyte

basophil

eosinophilic
promyelocyte

eosinophil

neutrophilic
promyelocyte

Neutrophil
(PMN)

myeloblast

mast
cell

Functions of Phagocytic Cells

Macrophages and Neutrophils

Localization and removal of foreign substances

(ie., pathogens) (MP and PMN)
Inflammation- response to infection or injury
(MP and PMN)
Initiate wound healing (MP)
Secretory functions (MP)
Immunologic functions: process and present
antigen (MP)
Tumor cell surveillance and cytotoxicity (MP)

Functions of Phagocytic Leukocytes

(mono, MP and PMN)

Localization and Removal of Foreign

Substances
(Inflammation and wound
healing)
-Chemotaxis: directed migration to site of injury;
chemical mediators (chemotactic factors)
-Phagocytosis: ingestion of foreign substances
-Metabolic destruction: digestion and killing
Oxygen-dependent (MPO, ROI, RNI)
Oxygen-independent (cationic proteins,
lysozyme, TNF, porphorins

Secretory Functions of
Macrophages
Second most potent secretory cell in
body

Enzymes capable of degrading

extracellular matrix proteins (collagenase,
elastase)
Products involved in host defense (ROI,
RNI, eicosanoids)
Regulatory proteins (IL-1, TNF, pGE2)

Macrophage-derived Mediators

Binding proteins (transferrin, fibronectin)

Complement components
Proteolytic enzymes (lysozyme, neutral protease, acid
hydrolyases)
Enzyme inhibitors (2-macroglobulin plasmin inhibitor)
Endogenous pyrogen (IL-1)
Reactive oxygen intermediates (superoxide, hydrogen
peroxide, hydroxyl radical)
Reactive nitrogen intermediates (nitric oxide, peroxynitrite)
Bioactive lipids (PAF, PG, LT, TBX)
Chemotactic factors
Growth factors/cytokines (IL-1, TNF, IL-6, FGF, CSF)

Macrophage Mediators Can also

Damage Host Tissues
Bioactive
Lipids

ROI
Activated
Macrophage

TNF-
IL-1
chemokines

Proteolytic
Enzymes

RNI

Production of ROI
(Respiratory Burst)
Requires NADPH oxidase
NADPH NADP+
+
2 O2

2 O-

Superoxide anion

Superoxide dismutase

2 O-

Hydrogen peroxide

H 2 O2

Myeloperoxidase (PMN)

H2O2 + C1-

Chloramines

Reactive Oxygen Intermediates

H2O2
O2OH-

Lipid Peroxidation
Membrane, Protein and DNA
Damage

Reactive Nitrogen Intermediates

Nitric oxide and peroxynitrite

Nitric oxide- formed from l-arginine by the
enzyme nitric oxide synthase (NOS)
Macrophages: (NOSII) induced by
inflammatory cytokines (IFN, TNF) and
bacterially-derived products (LPS)
Highly labile; oxidizes nucleic acids,
membranes, proteins
Nitric oxide reacts with superoxide anion
forming peroxynitrite
Antitumor/antibacterial activity

Proinflammatory Cytokines
Tumor necrosis factor-
Interleukin-1
Interleukin-6
Interleukin-18
Chemokines
Interferon-

Tumor Necrosis Factor-

Proinflammatory
Primes

phagocytes to produce
ROI and RNI
Cytotoxic
Induces apoptosis and necrosis

Immune Functions of Macrophages

Tumor Surveillance

Tumor Cytotoxicity

Antigen processing and

presentation

Immune Functions of Macrophages

Antigen processing and

presentation

Tumor Cytotoxicity

Tumor Surveillance

Macrophage Processing of
Antigens

Macrophages function as accessory cells or

antigen processing cells
Macrophage associated antigen is 1000x more
immunogenic
Processing of antigens involves change so that it
binds MHC II (Ia) proteins; may involve
unfolding, partial degradation, selection for
epitope with high affinity for MHC II
Required for T-helper cell recognition of antigens
Other APC: B cells, epithelial cells

Antigen Processing
Phagocytosis of antigen
Partial degradation or unfolding
Binding to MHC II (Ia) proteins
Re-expression of processed antigen
on cell surface
Presentation to T-helper cells

Antigen Processing and Presentation

II

Antigen
presentation

Macrophage (APC)
MHC
Class II

CD4

Processed
Antigen
T cell receptor

Cytokines

CD4 Th Cell

Adaptive Immunity:
Humoral Immunity
Mediated by B lymphocytes which produce
antibodies or immunoglobulins (Ig) in
response to antigen challenge
Antibodies: glycoproteins; selective, highly
specific; found in globulin fraction of
serum (humoral=blood)
Five Classes: physical, chemical and
antigenic differences

Classes of Antibodies
IgM: primary immune response (7%); Type III

hypersensitivity reaction; immune complexes;

B cell receptor
IgG: secondary immune response, B memory
cells (70%)
IgA: external secretions, produced locally against
bacteria and viruses (15%)
IgE: Type I hypersensitivity reactions, minute
amounts
IgD: umbilical cord blood, primitive recognition or
regulation; B cell receptor

Antibody Structure
Fab
Light chain

N-terminus
ss

ss

Heavy chain
C-terminus

Fab Variable region

antigen binding site
papain (2 Fab fragments)

-ss-

Fc

pepsin ( 1 Fab fragment)

Cellular binding site

Interchain disulfide bonds stabilize domain structure

which is tied to Ig function; differences in Ig
molecular weight due to differences in heavy chains

Antibody Structure
IgM: pentamer

YY

IgA: monomeric; becomes multimeric in

endothelium; acquires glycoprotein that protects
it against digestion

Kinetics of Antibody
Production
Primary immune response: IgM
Secondary immune response : IgM and IgG

Shorter lag
Higher levels of specific IgG produced
Steady state level persists longer
IgG predominates
Quantitative difference between primary and
secondary immune response due to increase in the
number of potentially reactive B cells
Adjuvants: IgG produced after primary response;
secondary IgG response sustained

Kinetics of Antibody Production

B Lymphocyte Differentiation
Bursa
equivalent

B cell

Pre-B
cell

Antigen

bone
marrow

10%

90%

T cellprocessed
antigen

Y
Sensitized B cell
Plasma cell

Memory cell

MP

complement

Antigen

AG

Lysis

Immune Response: B Cells

Maturation/
Differentiation

Large
B cells

lymphoblast

Y
Antigen

U
lymphoblast

B
B
memory

Small
B cell

Large
B cells

B
memory

B
B

small
B cell

Clonal
Proliferation

Antibody
triggering

Monoclonal Antibodies
Technique developed by Miller and Kohler (1980);
revolutionized immunology; involves development
of single antibody secreting immortalized cell
Inject mice with antigen (2 x)
After 2 weeks, collect spleen cells (B cells);
HGPRT+, Ig+, G Fuse with mouse myeloma (HGPRT-, Ig-, G+) in PEG
and HAT (hypoxanthine, aminopertin, thymidine) medium
Collect and clone hybridoma cells (HGPRT+, Ig+, G+)
Select for clones that produce specific antibody

Monoclonal Antibodies

Monospecific: normally antibodies extremely

polymorphic even when directed against single
antigen; normal antisera heterogeneity, therefore no
two antisera identical

Reproducible: identical antibody more specific

and reliable

Unlimited quantities: permanent cell line, grows

indefinitely and produces very large amounts of
antibody.

Antigen need not be pure or characterized:

antibody reacts with only one determinant

Monoclonal Antibodies
Potential Use

Reverse biological effects of certain agents

(digitalis, morphine, kepone, histamine)
Drug scavengers: low levels drugs and
chemicals (dioxin)
Drug targeting: cancer treatment
Tumor targeting
Identification and purification of proteins

Antigens

Substance recognized as foreign

Hapten: foreign substance that binds antibody;
does not elicit immune response
Immunogen: foreign substance that binds antigen
and elicits immune response
Carrier: large protein
Hapten + Carrier = Immunogen
Epitope: part of antigen that is recognized by
antibody
Immunodominant site: epitopes that are more
highly charged or more accessible

Types of Antigens

T-independent antigens
Complex carbohydrates
Do not require processing
Can directly interact with B cells
No memory

T-dependent antigens
Require macrophages or other APC
Require T-helper cells
Require major histocompatibility antigens
Mostly proteins

Antibody-Antigen Interactions
Binding of antigen to antibody
Occurs in variable region of antibody
molecule
Instantaneous
Exothermic
May form complexes
Cytotoxicity mediated by complement
(lysis)

Cell Mediated Immunity

Mediated by T lymphocytes which release

soluble mediators (lymphokines)

Important in host defense against viruses,

certain bacteria, fungi, transplant rejection and
tumor surveillance; Type IV (delayed type)
hypersensitivity (DTH) reactions

T Cells: derived from precursor cells in bone

marrow; mature in thymus; become educated

Types of T Cells

Regulatory T cells
T-helper cells: T4 (CD4) antigens (Th1, Th2)
T-regulatory (suppressor cells): surface
CD4+/CD25+; intracellular FoxP3 (naturally
occuring Treg)
Cytotoxic (killer) T cells: CD8 antigens
Identified by surface markers (CD4, CD8, CD25)
(monoclonal antibodies)
Work together to modulate immune response
Actions mediated by cytokines

T-helper cells
TH1 cells: T helper inflammatory cells

Activate macrophages
Involved in cell-mediated allergies, e.g. poison ivy
Promote rejection of transplanted tissue
Stimulated by cell-bound antigen, and secrete
lymphokines

TH2 cells: Stimulate B cells to produce

antibodies

T Cell Lymphokines
Macrophage activity factor (IFN
Macrophage inhibitory factor (MIF)
Interferon (IFN and IFN)
Lymphotoxic factor
Interleukin 2 (IL-2,T cell growth
factor)
Interleukin 3 (IL-3, B cell growth
factor)

T Lymphocyte Differentiation
Bone
marrow

education

Thymus

MP
MP

Th
(+)

(-)

PA
(+)

Sensitized
Th

Treg

(+)
Pre-T

(-)
Tcyt

cytokines

(-) B cell
(+)
Cytotoxic
lymphokines

T Cell Education

Learning to distinguish between self and nonself

Controlled by MHC (self marker) proteins
Selection of subpopulations of immature T cells
through their interactions with MHC proteins
displayed to them in thymus
Immature T cells recognize MHC alone; as they
mature, recognize MHC+antigen
Maturation dependent genetic alteration in T
cell receptor
Uneducated T cells are destroyed in thymus

Antigen Recognition by T Cells

Specific T cell receptor

T cells are MHC restricted; only recognize
antigen and MHC protein
T-helper cells recognize processed antigen
and MHC II (self)
Cytotoxic T cells recognize processed
antigen and MHC I (non-self or altered self)

Major Histocompatibility Complex

(MHC)
Large cluster of immune response genes; code for
proteins that direct T cell responses (mouse chrom 6;
human chrom 17); MHC I and MHC II
Allow immune system to distinguish self and non-self; to
destroy non-self or altered self (virally infected cells
and tumor cells)
Extreme polymorphism: millions of alleles or variants;
control individual responsiveness to antigen; inherit
haplotypes
Every individual unique; MHC markers genetically
determined; the closer two individual are genetically,
the closer their haplotype

Major Histocompatibility
Complex
Class I MHC: expressed on all somatic
cells; classic transplantation antigens

Class II MHC: expressed on immune cells

(B,T, macrophages, thymus epithelium);
immune associated (IA) antigens; important in
immune regulation, cell-cell communication

Class III MHC: complement

Activation of T Cells
altered
self

Macrophage

Antigen

T cell
receptor

CD4+

MHC II

processed
antigen

epitope

CD8+

T-helper

T-killer
Stimulatory cytokines
IL-3
B

IFN
MP

IL-2
T

Cytotoxic lymphokines

Potential Effects of Toxicants on

the Immune System

Immune Suppression
Decreased resistance to infection, impaired
surveillance; increased incidence cancer; delayed or
aberrant wound healing
General immune suppressants
Pharmaceuticals: corticosteroids (macrophage inhibitors;
lympholytic); methotrexate, cyclophosphamide (cytotoxic
agents; bone marrow suppressants)
Environmental/industrial chemicals (solvents, PAH, benzene,
pesticides, heavy metals, some air pollutants)
Agents of abuse (caabinoids, ethanol, tobacco)
Physical agents (UV, ionizing radiation)

Potential Effects of Toxicants on

the Immune System

Immune Enhancement
Autoimmunity: immune response against hosts own
tissues and cells; autoantibodies produced (ANA);
joints, connective tissue most sensitive; failure of
organism to recognize its own cells as self; leads to
inappropriate immune response against self antigens
Genetic factors
Chemicals/drugs
-lupus:
lupus genetic predisposition vs drug induced (hydralazine,
procainamide); increase Th or decreased Treg/s cells
-acute liver failure: alterations in self antigens; tienilic acid,
triglitizone

Potential Effects of Toxicants on

the Immune System

Immune Enhancement
Hypersensitivity/allergic reactions: most common toxic
manifestations of chemicals; undesirable reactions produced by
normal immune system; require presensitization; most common in
lung (asthma-dust; pneumonitis- inorganic chem.; DTHberyllium), skin (contact dermatis) and GI track; drugs/chemicals
can act as haptens by conjugating to proteins or cells; become
immunogens; ex, penicillin allergy; poison ivy
Chronic Inflammation:
Inflammation aberrant wound repair; foreign body
reaction; granuloma
General Immune Stimulation: nonspecific stimulation
macrophages and/or lymphocytes; ex, levamisole, copolymer,
estrogen, somatotropin

Hypersensitivity Reactions
Type I: immediate (anaphylactic) reaction (within 15-30 min);

can involve skin (urticaria, eczema), eyes (conjunctivitis),

nasopharynx (rhinitis), bronchopulmonary tissue (asthma) or
GI (gastroenteritis)

Primary exposure to antigen: binding of IgE to mast cells

Secondary exposure: cross bridging of antigen to IgE, capping,
Ca++ influx, release of vasoactive autacoids (ex., histamine,
LT, PAF)
Response: Inhibition of vascular smooth muscle; bronchiole
constriction, hay fever, uticaria, acute anaphylaxis
Examples: latex, peanuts, isocyanates, cobalt, nickel

Mast Cell Mediators

Histamine (increased vascular
permeability, constrict smooth muscles)
Serotonin (Increase vascular
permeability)
Eosinophil chemotactic factor
SRS-A (LTB) (constrict smooth muscles;
increased vascular permeability)
Platelet activating factor (platelet
aggregation)

Hypersensitivity Reactions
Type II: Cytotoxic hypersensitivity; antibody
dependent (IgG and IgM)

Binding of antibody to antigen

Complement fixation
Activation of complement
Lysis of antigen/antibody complex by complement
Phagocytosis and degradation by macrophages
Examples: blood transfusions, organ transplants;
hemolytic anemia; Rh disease; Farmers lungbiological dusts (hypersensitivity pneumonitis)

Hypersensitivity Reactions
Type III:

Intermediate reaction (6-24 hr)

Immune complex hypersensitivity; (Ab-Ag complexes)
Formation of immune complexes; mostly IgM; circulate in
blood; attach to vessel walls (esp. kidney); complement
fixation, phagocyte and platelet accumulation; increased
vessel permeability, edema, decreased blood flow, release
of hydrolytic enzymes and vasoactive substances; vessel
rupture, hemorrhage
Examples: arthus skin reaction-local; serum sicknesssystemic, protracted anaphylaxis, occupational diseases

Hypersensitivity Reactions
Type IV:

Delayed Type Hypersensitivity (DTH)

Reactions (48-72 hr)
Mediated by macrophages (MP) and cytotoxic T cells
(Tcyt)
Macrophage present processed antigen to T cells;
activation of Tcyt; MP and Tcyt release cytotoxic
mediators (CC, IL-2, IFN, TNF);
Examples: Tuberculin type allergy- other allergies of
infections; autoimmune diseases; allergic contact
dermatitis (acids, resins, preservatives, cosmetics,
beryllium, chromium, nickel)- agents act as small
MW haptens; form complete antigens by binding to
dermal proteins.

Proceedures for Assessing

Immunotoxicology

Immunopathology
Good for assessing general immune alterations
-Hematology profile: leukocyte counts, differentials
-Clinical chemistry: Ig levels
-Histology: BM, nodes, spleen, thymus
-Organ weights: spleen, thymus

Host Resistance Assays

potentially useful as initial screen to measure
immunocompetence; rationale- immunosuppressed hosts are more susceptible to
infections or to cancer

Host Resistance Assays

Tumor Cell Challenge

Measure resistance to tumor cell graft
following toxicant exposure
Inject tumor cell suspension into flank;
palpate weekly for tumor development;
measure latency and volume
Examples: Lewis lung carcinoma (DES);
MCA sarcoma (PCP); polyoma virus (TCDD,
BP, urethane)

Host Resistance Assays

Infectious Agents
Range of effects may be observed: mild
infection, severe infection, death
Injection of infectious agent can lead to
activation of macrophages and T cells which
decrease severity of disease or to general
immune suppression and increased severity of
disease
Monitor effects of toxicant on frequency and
latency of infection/mortality and effects on
specific target organs

Examples of Chemicals and Drugs that

Alter Host Resistance to Infections

Viruses
HSV (PCB)
Influenza (DTT)

Bacteria
E-coli (lead, cadmium)
Salmonella (lead, TCDD, PCB)
Lysteria (TCDD, DES, cyclophosphamide)

Parasites
Trichinella (PCB)

Assessment of Phagocytic Cell

Functioning
Differential counts
Measurement of functional activity
(chemotaxis, phagocytosis,
cytotoxicity-bacteria, tumor cells)
Lysosomal enzyme activity
Clearance capcity RES (colloidal
carbon, 125-I-triolein)

Assessment of Humoral Immunity

Measurement of antibody titers (immunodiffusion,

hemagglutination, RIA, ELISA)
Measurement of antibody synthesis (plaque
forming assays)
Measurement of B cell surface receptors (EAC
rosettes)
Enumeration of B cells (Mab)
Measurement of B cell proliferation (polyclonal
activators-LPS, PWM)

Assessment of Cell Mediated

Immunity

Enumeration of T cells (Mab)

Measurement of T cell proliferation in response
to mitogens (PHA, Con A)
Assessment of DTH response: measure skin
response to sensitizer (oxasalone, DCNB)
Graft vs Host reaction
T cell cytotoxicity- tumor cells
Lymphokine production
Mixed lymphocyte reaction