609 1st int slides
TRANSCRIPT
-
8/6/2019 609 1st Int Slides
1/34
IMMUNOLOGY
-
8/6/2019 609 1st Int Slides
2/34
What does the immune system recognize? Most immunogens
contain multiple antigenic determinants called epitopes.Anepitope is the basic minimal structure on the Ag surface
recognized by the immune system.Most immunogens are
composed of a variety of epitopes, haptens comprise a single
epitope.Paratope is the corresponding specific binding site
on the Ab molecule
Antibodies with a complimentary shape will be formed that
can specifically combine with these epitopes on the antigen.
one Ag may stimulate the prod. of many Abs but only the bestfit will combine
-
8/6/2019 609 1st Int Slides
3/34
Antigen-Antibody binding
-
8/6/2019 609 1st Int Slides
4/34
ANTIBODY
REFERS TO A GRPOFSERUM GLYCOPROTEINS,FORMEDIN
RESPONSETO ANANTIGEN. ANTIBODIESFOUNDINSERUMCALLED HUMORAL ABS
BROADLY KNOWNASIMMUNOGLOBULINS ---BELONG
TO FIVECLASSES - IgG,IgM,IgD,IgA,IgE
MAYBEDESIGNATEDBYNAMES THATDESCRIBETHEIRREACTIONWITH CERTAINTYPES OFANTIGENS
ANTITOXINS (NEUTRALIZEMICROBIAL TOXINS),
AGGLUTININS(CAUSECLUMPING),PRECIPITINS(CAUSEPRECIPITATION),LYSINS(CAUSELYSES) COMPLEMENTFIXING
ANTIBODIES(CAUSES CFTREAC.),OPSONINS(MAKEMICROBESMORESENSITIVETO PHAGOCYTOSIS)
-
8/6/2019 609 1st Int Slides
5/34
Immunoglobulins & functions
There are 5 classes ofIg
class Mol.wt % in serum function
IgG 150,000 80 Involved in phagocytosis
CFT, found in lymph/serum,
crosses placental barrier
IgM 900,000 6 Found in serum & lymph, cidal
to Gm-ve bacteria
IgA 170,000 13 Found in tears,saliva otherbody secretions
IgD 185,000 1 Found in serum
&lymphocytes, contr B cell
stim
IgE 190.000 0.002% Causes allergies, Imm hyper
-
8/6/2019 609 1st Int Slides
6/34
Immunoglobulin structure Each Ig mol is made of 4 polypeptide chains :
2 identical light(short) & 2 identcal heavy(longer) heldtogether by interandintra-disulphide(S-S) bonds There are 2types of light chains-kappa & lamda
Basedon this the Ig mol may be kappa/lamda
In order to study the diff.portions of Ig mol, it was treatedwith
proteolytic enzymes-papain & pepsin.
Papain digestion results in3 pcs----2 identical pcsFab(fractionAg binding) each containing a light chain andhalfa heavychain. The 3rdsingle pcFc(fraction-crystallisable) consists of
the other 2 halves of the heavy chain still linkedby thedisulphide bonds
When Ig is digestedwith pepsin only one major fragment of 2Fab units is obtainedwhere the interdisulphide bondis stillexisting andan Fc region which loses its link& breaks up
-
8/6/2019 609 1st Int Slides
7/34
Structure of
Immunoglobulin
-
8/6/2019 609 1st Int Slides
8/34
Papain digestion results in 3 pieces-2 identical pcs (Fab) each containing a light chain
and half a heavy chain,the 3rd pc (Fc) consists of other 2 halves of the heavy chains
still linked at the hinge by the disulphide bonds
-
8/6/2019 609 1st Int Slides
9/34
When IgG is digested with pepsin, only one major fragment consisting of two Fab
Units is got, it is joined at the hinge with interdisulphide bonds. The Fc region loses
Its link with the hinge region and breaks into small fragments
-
8/6/2019 609 1st Int Slides
10/34
Based On cellular basis immunity may be of 2 types
Humoral Immunity Cell mediated immunity
Involves syn.& release the antigen stims
of free Ab into blood / the produc. Of sensi-
other body fluids tized lymphocytes which
Depend on B cells confer Host protection
depend on T cells
Cells involved in immunity are present throughout the body but
mainly found in Lymphoreticular organs
a) Primary lymphoid organs---thymus & bursa of fabricus
b) Secondary lymphoid organs---lymph nodes, spleen, tonsils
-
8/6/2019 609 1st Int Slides
11/34
Active immunityActive immunity is the resistance developed by an individual in
response to an antigenic stimulus. The antigen may gainentrance either by natural infection or through vaccination(artificial). Active immunity involves the active functioning of apersons immune system, resulting in the synthesis of specificantibodies (humoral immunity) or production of
immunologically active cells (cell mediated immunity)Humoral immunity-here specific antibodies are produced in in
plasma or lymph,in response to specific antigenic stimulus. Theantigen may then be clumped or their toxicity neutralized.Thisimmunity sets in after a latent/lag period needed for Ab prodn
Cell mediated immunity-here instead of specific antibodiessensitized lymphocytes are produced which react with thespecific antigens bringing about cytotoxic effects and lysis.If aperson is exposed to a parti. Ag for 2nd time, immune responseis quicker & abundant than 1st ---known as Secondary response
-
8/6/2019 609 1st Int Slides
12/34
-
8/6/2019 609 1st Int Slides
13/34
Immune system The cells involved in the immune responses have their origin in
the stem cells of bone marrow.
There are 2 types of cells inthe bone marrow
Myeloid lineage---neutrophil, monocytes,basophil, esonophil
Lymphoid---B lymphocytes & T lymphocytes
M
ajority of lymphoid stem cells migrate to the thymus gland &get differentiated into T cells ofHelper,Suppresor& Cytotoxic
type. This starts shortly before birth, they are small, found in
blood, lymph, lymphoid tissue. T cells do not produce humoral
antibodies but indirectly control their production. T cells are
involved in cell mediated immunityOther lymphoid stem cells migrate to the Bursa(lymphoid area in
birds) equivalent eg payers patch & differentiate into B cells
when stimulated by antigen produce antibodies to be circulated
through out lymph & blood to protect the individual, lives for Ito 2 weeks.Involved in humoral immunit
-
8/6/2019 609 1st Int Slides
14/34
Primary lymphoid organs are the major site for lymphopoiesis. Here the
lymphoid Stem cells proliferate, differentiate and mature into immuno-
competent cells in the absence of antigenic stimulation.
The mature but nonfunctional lymphocytes migrate from the primary lymphoid
organs through the blood or lymph and accumulate in the peripheral or
secondary lymphoid organs.
Here the cells when exposed to antigenic stimulus become functional by
producing either 1) specific antibodies or 2) sensitized cells.
The spleen, lymph nodes, tonsils, appendix constitute the secondary lymphoid
organs. These second.Lymph.Orgs are poorly developed at birth and grow
progressively while the primary ones are large at birth and atrophies on age.
-
8/6/2019 609 1st Int Slides
15/34
IMMUNE RESPONSE
THE BODY CELLS IMPORTANT TO IMMUNE RESPONSE
B CELLSLYMPHOCYTES WHICH PRODUCE ABS. IN BLOOD
MACROPHAGES---PHAGOCYTIC CELLS WHICH ALERT HELPER T CELLS OF
PRESENCEOF PATHOGEN
HELPER T CELLS--- MASTER SWITCHES WHICH RELEASE LYMPHOKINES---STIMULATES RAPIDDIVISION OF T & B CELLS
NATURAL KILLER(NK) CELLS---LYMPHOCYTES THATRELEASELYMPHOTOXINS
DIRECTLYDESTROYBODYCELLS ALREADY INFECTED WITH PATHOGEN
SUPRESSOR T CELLS---LYMPHOCYTES WHICH SLOWS DOWN IMMUNE
RESPONSE BYINHIBITING THEPRODUCTION OF CYTOTOXIC T CELLSCHECKING THEM FROM CAUSING MORE DAMAGE THAN NECESSARY
MEMORY CELLS---A GROUP OF T&BCELLS PRODUCEDDURING PRIMARY
ENCOUNTERBUT ARERESERVED TORESPOND TOLATER ATTACKS BY SAME
ORGANISM (role in secondary immunity)
-
8/6/2019 609 1st Int Slides
16/34
Function of B cells & T cells
B cells have Ig receptors on their surface membrane which aresuitable for the specific Ab it will produce.If the B cells in the
lymph nodes encounter an antigen,which has been transported
to the lymph node via the lymphatics,the cells get
activated,divide and differentiate to become plasma cells
B cells must receive a signalfrom T cell/macrophage to
proliferate andonlyafteralag perioddifferentiate into plasma
cells in the bloodwhich produce the antibodies to fight.
The function of the T cells is to recognise an antigen andthen
release *lymphokines (helper T)---stim B & T cells to grow &
differentiate *enhance suppresor function ( supp T)---which
inhibit the production of killer T cells that actually secrete
lymphotoxins and bring about cell lysis
-
8/6/2019 609 1st Int Slides
17/34
Innate vs adaptive immunity
Innate system(non-specific, natural )Adaptive system( specific, acquired )
Does not take time to respond, as
the system is in place prior to
exposure to antigen
Takes some time to react to an
invading organism, since the system is
induced after exposure to antigen
It is not antigen specific andreacts
equally well to a variety of
organisms
It is antigen specific and reacts onlywith that particular organism
Does not demonstrate
immunologicalmemory
This system demonstrates
immunological memory. It remembers
that it had encountered an invading
Organism and reacts more rapidly on
subsequent exposure.Plays an
important role in secondary immunity
-
8/6/2019 609 1st Int Slides
18/34
Primary/secondary immune response
P
rimary immune responseimmune response produced byentry of the Ag for the first time---the 1st injection of the Ag is
called primary dose.
Secondary immune response if the same animal is exposed to
the same Ag for the second time the animal produces the
immune response for the 2nd time called secondary immuneresponse and the 2nd Ag is known as booster/secondary dose.
The humoral immunity results in the production of antibodies in
the blood. The amount of antibody produced is called the
antibody titre.If this is plotted against time a sigmoid curve isobtained having 4 phases lag, log, plateau and decline phases.
Graphical representation of primary/secondary immune
response
-
8/6/2019 609 1st Int Slides
19/34
comparisonPrimary immune response Secondary immune response
Lag phase- time after the Ag exposure
Varies from hours to days
Very short , the body responds quickly
Log phase rise in Ab content and this is
sharp
The Ab titre here exceeds that of primary
response ----10 folds more
Plateauthis is a time when the Ab titre
is constant, short duration
The time is much extended compared to
primary
Declinehere the catabolism of Ab
exceeds the production, so there is a fallin Ab titre
The fall in Ab level takes place very slowly
-
8/6/2019 609 1st Int Slides
20/34
Clonal selection theoryLong standing principle governing Ab production
Each B-lymphocyte during its development is committed to
respond to one antigenic determinant. (epitope)
Accordingly each lymphocyte has a single type ofAg-specific
receptoron its surface. (paratope)
Receptor occupation is required for B cell activation.
Following contact with Ag, the B cell expands and divides
producing aB cell clone ( clone is a genetically identical
progeny produced naturally/artificially by mitosis from a
single specific cell type). This will differentiate into plasmacells which will form antibodies having receptors of identical
specificityas the parent B cell
-
8/6/2019 609 1st Int Slides
21/34
IMMUNOTOLERANCE
ITIS A CONDITIONWHEREadministration OFANTIGENICMATERIAL DOESNOTEVOKEANIMMUNOLOGICAL RESPONSE
BOTHB& T CELLS AREMADEUNRESPONSIVE
immunotolerance
natural or self tolerance
Where body does not evoke
immune response to self
antigens
Induced tolerance
Where body does
Not evoke immune
Response to external
antigens
During pregnancy mothers tolerance to foetus/placenta ---eg of natural
In clinical practice during organ transplantation chances of rejection is major
Problem faced. To prevent rejection, drugs are used to induce tolerance
-
8/6/2019 609 1st Int Slides
22/34
Tolerance can occur in several ways -----
1) GENETIC UNRESPONSIVENESS-------ANIM AL LACKS THE GENETIC
ABILITY TO RECOGNIZE ANTIGENIC MATERIAL
2) TSUPRESSION------Supressor T CELLS MAY BE MORE EFFECTIVELY
ACTIVATED THANTHE helper T CELLS THUS SUPPRESSING THE
IMMUNESYSTEM
3) HELPLESSNESS----WHENT cells CANNOT ACTIVATETHEB CELLS THEY
AREREGARDED AS HELPLESS
4) CLONAL DELETION----CONTACT WITH ANTIGENINTHE NEONATE (
immature immune system) RESULTSINDEATH/PERMANENTINACTIVATION
OFdeveloping LYMPHOCYTES
-
8/6/2019 609 1st Int Slides
23/34
Autoimmunity
Antigens present in ones own cells are autoantigens or self
antigens. These generally do not elicit an immune response.
Only the foreign or nonself Ag evokes immune response.
However when they are altered by the exposure to some
bacteria/ toxic chemicals/drug, they are recognised as non-selfAgs & stim prod. of Abs against them called autoantibodies.
Hence autoimmunity is definedas humoral/cell mediated
immune response directedagainst the bodys own tissues or self
antigens
Autoimmune diseases are a group of disorders in which host
tissue damage is caused by autoimmune responses to self Ags
-
8/6/2019 609 1st Int Slides
24/34
Types of autoimmune(AI) disease
1. Haemolytic Auto Imm diseaseresults in destruction of
RBC/platelets/ as Ags on RBC etc become non- self Ag.
Haemolytic anaemia, leucopenia,thrombocytopenia (low
platelet count )2.Localised Auto Imm disease results in the destruction of a
particular organ eg Thyrotoxicosis, Addisons disease
Myasthenia gravis (weakening of skeletal muscles-autoAb
against Ach receptor cells thus Ach cannot be producedno
impulse passed)
3.Systemic Auto Imm disease affects the whole body or many
organs
eg Lupus erythematosus, rheumatoid arthritis
-
8/6/2019 609 1st Int Slides
25/34
Hypersensitivity Immune response is mostly directed towards the protection
of host, as the bacteria/virus is killed in the process. Howevernot all Ag-Ab reactions are beneficial.
In hypersensitivity the immune responses become
injurious/destructive to the host, killing or damaging body
cells Thus it may be definedas the violentadverse reaction
(malfunction) of the immune system leading to severe
symptoms & even death in a sensitizedanimal/human when it
is re-exposedto the same Ag for the 2nd time
Hypersensitivity occurs only in 10% of cases, in clinical terms
The agents causing hypty are called allergens
The effect is called allergy/hyperty
-
8/6/2019 609 1st Int Slides
26/34
Factors causing hypersensitivityAllergens may be extrinsic (enters into body) or intrinsic
(present inside body)Drugs penicillin, sulphacetamide, aspirin
Airborne particles pollen grains, dust, scales/feathers ofanimals eg dog, cat, bird
Food stuff shell fish, brinjal, pineappleInfectious organisms bacteria, virus, fungi, parasite
Blood transfusion mismatched blood
Common hypersensitive reactions---
Anaphylaxis Serum sickness
Transfusion reaction Mantoux reaction
Erythroblastosis foetalis Contact dermatitis
Arthus reaction Graves disease
-
8/6/2019 609 1st Int Slides
27/34
Classification
Based on time taken-*immediatesymptoms appear eg Hay fever within mins i.e
rapid onset (type 1, 2, 3), eg allergic reacts-urticaria (skin),
rhinitis, extrinsic asthma,involves B cells, can be transferred
through serum, controlled by antihistamines
*delayed symptoms within 24 to 48 hrs (type 4,5) eg Mantoux
test, involves T cells, transferred through T cells/corticosteroids
Based on different mech. of action
*type 1 :Anaphylactic hypersensitivity
*type ii:Antibody-dependent cytotoxic hyperty
*type iii:Immune-complex mediated hyperty
*type iv:Cell mediated hyperty
*type v: Stimulatory hyperty
-
8/6/2019 609 1st Int Slides
28/34
Typescontd.Anaphylaxisis defined as exaggerated reactions shown by the
body to a foreign substance to which it has previously become
sensitized.
Abs are fixed on mast cells in host causing release ofhistamine, serotonin on 2nd exposure eg penicillin allergy
Cytotoxic when antibodies attach to the antigens located on the
surface of cells eg agglutination & lysis occurs due to
mismatched blood groups, eg erythroblastosis ( RBC rupture)
Immune complexwhen huge amt of antigen enters body a large
conc. of Ab is produced which combine to form insol complex
which get attached around minute blood vessels leading to
hypersensitivity, Arthus reaction, serum sickness
Cell mediatedcaused by interaction between Ag & T cells, abs arenot involved Contact dermatitis, tuberculin reaction
Stimulatorycaused by interaction between Ab with cell surface
Ag where over stimulation of cells of the thyroid occur eg
Graves disease (thyrotoxicosis)
-
8/6/2019 609 1st Int Slides
29/34
ANTIGEN-ANTIBODY REACTIONSSEROLOGYIS THESTUDY OFAG-ABREACTIONS
USED------ TO TYPEBLOOD,DIAGNOSEDISEASE
IDENTIFYMICROBES,INTISSUETRANSPLANTATION, TO DETECT
ALLERGY/HYPERSENSITIVITY
THECOMMON Ag-Ab reactions (INVITRO TESTS)ARE---
AGGLUTINATION
PRECIPITATION
COMPLEMENTFIXATION
FLUORESCENT ANTIBODY TECHNIQUE
RADIOIMMUNOASSAY (RIA)ENZYMELINKEDIMMUNOSORBENT ASSAY(ELISA)
PURPOSE:THEPRESENCEOFSPECIFIC ANTIBODIESINTHEPATIENTSSERUM
WILL INDICATEWHETHER HEHAS THEDISEASE/HEISIMMUNETO
DISEASE/HEIS ALLERGIC TO ANTIGEN
-
8/6/2019 609 1st Int Slides
30/34
Antigen antibody reactionWhen an Ab and the Ag responsible for its production are mixed, the two
bind to form a Ag-Ab complex and such a reaction is called Ag-Ab react,represented by Ag + Ab ----------------- Ag-Ab complex
Characteristics of such a reaction:
1. Role -This immune complex forms the basis of humoral immunity
2. Specificityrefers to the indiscriminate ability of a particular Ab tocombine with only one Ag determinant responsible for its production.
Compared to lock and key system.
3. Binding sites of Ag and Ab. An Ag may contain 10-50 determinants
while most Abs are bivalent (2 paratopes) only IgM has 5-10
4. Binding forcesof Ag and Ab. The intermol forces are of non covalenttype eg hydrogen bonding/vanderwaal. Affinity refers to strength of bond
between a Ab mol and a single antigenic determinant
5. Avidity- It is the total strength of the bonds after formation of the
individual complexes on an Ag nAb + mAg--------------AbnAgm
n=no of Ab m=no of Ag determinants
-
8/6/2019 609 1st Int Slides
31/34
AGGLUTINATION --- is an antigen antibody reaction where the specific antibody
causes the insoluble cellular (particulate) antigens to adhere to one another to form
visible clumps,in presence of electrolyte and suitable temp, pH.
The particulate Ag include RBC, platelets, lymphocytes, bacteria, viruses etc.
Agglutination is used for ABO and Rh blood typing --- sample( A ,B, AB, O)
agglutination anti B
here the Abs are called agglutinins & Ags called agglutinogens
Applications----
Widal Test ---used for diagnosis of typhoid fever. The test uses antigen H & O, so if
the antiserum of patient carries specific Abs to these Ags, clumping seen.Well Felix reaction ---- used to test typhus fever
Coombs test---- devised for detection of anti Rh antibodies
sample+ +
Anti Aserum
AntiBserum
-
8/6/2019 609 1st Int Slides
32/34
Agglutination
Qualitativedone only to detect the presence of
Specific Ag or Ab in sample.
Eg patients blood can be mixed with serum having
anti A, B, AB, to see with which it agglutinates.
Grp A if agglutination with anti A serum(Ab in GrpB)
Quantitativedone to measure the level of AbsPresent in the serum with respect to Ag. Serial
dilutions of a serum sample is made and placed in
same vol in a 96 well plate.It is
Mixed with a fixed no. of the Ag(bacteria/RBC)
The max dilution that gives visible agglutination is
noted and is referred to as Titre.
A 4 fold rise intitre is generally taken as disease established.
Passive agglutination since agglutination takes place only with insoluble Ags
one modification can be done incase of soluble Ags (viral Ag, polysaca) the RBC is coated
with the sol Ag and the test is done as above. Hence by observing the clumping of RBC the
titre of the sol, Ab may be measured indirectly hence passive.
-
8/6/2019 609 1st Int Slides
33/34
Complement fixation test Complement refers to a group oflarge, thermolabile,complex
proteins(beta globulins),found in the serum and body fluids,
which completes the Ag-Ab complex, by fixing with it.
CFT is an immunological technique used for the detection of
unknown Ag or Ab by finding if the binding/fixing of the
complement to the Ag-Ab complex has occurred or not. When a complement is added to a serum containing Ag and
its Ab, the complement present is activated and immediately
binds to the Ag-Ab complex and is said to be fixed
Ag + Ab + * ======== Ag--*-Ab
compl fixed
-
8/6/2019 609 1st Int Slides
34/34
CFTThis test involves the operation of 2 immune systems
1st system ---is the unknown system under detection called testsystem
2nd system---is the known system containing sensitized RBC andacts as the indicator system as it indicates complementactivity
Steps in CFT
1.Serum to be tested for Ab + known Ag + compl = Ag-*--Ab
2. RBC (coated with its Ab ) is added to above = no haemolysisin the tube due to unavailability of free complement as it has
been used up in the 1st
complexResult : Absence of haemolysis is a +ve test indicating presence
of specific Ab in sample.Presence of haemolysis is ave test,indicating absence of specific Ab in sample