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Bio 151 lectures for examination 2 Bio 151 lectures for examination 2 Presentation Transcript

  • VACCINES REFERENCES: a) Abbas; b) Kuby; and c) Lectures of Dr Nina Gloriani, MD, PhDMonday, September 10, 2012
  • EDWARD JENNER • Father of Vaccinology & Founder of Immunology • Major contribution: first reliable method of conferring lasting immunity to a major contagious disease (smallpox) • Initiated the technique of vaccinationMonday, September 10, 2012
  • LOOKING BACK... • 18th century: Small pox • 19th century: Rabies; typhoid; cholera; plague • Early 20th century: BCG;Yellow fever; pertussis: influenza; diphtheria; tetanus • After world war II: • OPV; MMR; Adenovirus; Salmonella Ty21a; varicella; Injected Polio; Rabies; Japanese encephalitis; Hepatitis A; Pneumococcus; meningococcus, Hemophilus influenza PRP, Hepatitis B; Acellular pertussisMonday, September 10, 2012
  • RECALL: IMMUNITYMonday, September 10, 2012
  • ACQUIRING PASSIVE AND ACTIVE IMMUNITYMonday, September 10, 2012
  • ACTIVE & PASSIVE IMMUNITYMonday, September 10, 2012
  • PASSIVE IMMUNIZATION • Administration of preformed antibodies to treat infection • Can be life-saving where toxins are already circulating : tetanus, diphtheria, snake bite • Important where high titer specific antibody is requiredMonday, September 10, 2012
  • PASSIVE IMMUNIZATION • Preformed antibodies generally made in horses, but occasionally obtained from recovered patients • Over 1000 donors used for each pool of ISG, and sera are screened for HIV and hepatitis B and CMonday, September 10, 2012
  • Monday, September 10, 2012
  • COMMONLY USED AGENTS DISEASE AGENT Black widow spider bite Horse antitoxin Botulism Horse antitoxin Diphtheria Horse antitoxin Hepatitis A & B Pooled human IgG Measles Pooled human IgG Rabies Pooled human IgG Snake bite Horse antivenin Tetanus Pooled human IgG or horse antitoxinMonday, September 10, 2012
  • PASSIVE IMMUNIZATIONMonday, September 10, 2012
  • NON-SPECIFIC IMMUNOTHERAPY • Same compounds that act as adjuvants for vaccines also used on their own to boost the general level of immune activityMonday, September 10, 2012
  • IMMUNOTHERAPY OF TUMORSMonday, September 10, 2012
  • Monday, September 10, 2012
  • Monday, September 10, 2012
  • ACTIVE IMMUNITY • The process of administering antigen to a live host to induce an immune response for either academic or public health reasons • Vaccines have been developed as a prophylactic measure to prevent disease caused by infectious agents, provided their use caused only low levels of morbidity • WORLD HEALTH ORGANIZATION: Expanded Program on Immunization (EPI) and Vaccine development programs/initiativesMonday, September 10, 2012
  • GENERAL TYPE OF VACCINESMonday, September 10, 2012
  • ANTIGENIC PREPARATIONS USED AS VACCINESMonday, September 10, 2012
  • COMPARING NOTESMonday, September 10, 2012
  • LIVE ATTENUATED VACCINESMonday, September 10, 2012
  • EFFECT OF VACCINES ON VIRAL DISEASESMonday, September 10, 2012
  • KILLED-WHOLE ORGANISM VACCINEMonday, September 10, 2012
  • POLIO: LIVE VERSUS KILLED VACCINEMonday, September 10, 2012
  • Monday, September 10, 2012
  • SUBCELLULAR FRAGMENTS VACCINESMonday, September 10, 2012
  • Monday, September 10, 2012
  • TOXIN-BASED VACCINESMonday, September 10, 2012
  • Monday, September 10, 2012
  • DNA VACCINESMonday, September 10, 2012
  • THE MALARIA VACCINE STRATEGYMonday, September 10, 2012
  • VACCINES IN GENERAL USEMonday, September 10, 2012
  • VACCINES RESTRICTED TO CERTAIN GROUPSMonday, September 10, 2012
  • SAFETY CONCERNS WITH VACCINESMonday, September 10, 2012
  • WANTED VACCINES...Monday, September 10, 2012
  • ADJUVANTSMonday, September 10, 2012
  • EFFECTS OF ADJUVANTS • Concentration of antigen in a site where lymphocytes are exposed to it • the “DEPOT” effect • Induction of cytokines which regulate lymphocyte function • cytokines themselves shown to be effective adjuvants, particularly when coupled directly to antigenMonday, September 10, 2012
  • START HEREMonday, September 10, 2012
  • VACCINE CONSIDERATIONS #1 • Type • Indications • Target group • Dose • Route of Administration • Monitoring Side EffectsMonday, September 10, 2012
  • TARGET GROUPSMonday, September 10, 2012
  • Monday, September 10, 2012
  • MONITORING • Side effects: Fever • LOCAL SYMPTOMS AT INJECTION SITE: Soreness, redness, swelling size • GENERAL symptoms– headache, malaise, loss of appetite, nausea, vomiting • Adverse Reactions • ContraindicationsMonday, September 10, 2012
  • VACCINE CONSIDERATIONS #2 • SAFETY: Toxicity, teratogenicity, pyrogenicity, adverse reactions • IMMUNOGENICITY: Seroconversion: levels of antibody before and after immunization; Geometric mean titer (GMT): antibody titer following vaccination • EFFICACY: correlation between seroconversion and PROTECTION (Antibodies; CMI or both)Monday, September 10, 2012
  • SAFETY: ADVERSE REACTIONS • MILD: adverse reaction does not interfere with normal activities; may be bothersome • MODERATE: adverse reaction produces some impairment of functioning but NOT hazardous to health; uncomfortable or disturbing • SEVERE: adverse reaction produces significant impairment of functioning or incapacitation; a definite hazard to healthMonday, September 10, 2012
  • EFFICACY • Streptococcus pneumoniae; Neisseria meningitidis; Hemophilus influenzae capsular polysaccharide (85%) • Diphtheria and tetanus Toxoids (>90%) • Bordetella pertussis (whooping cough) (>90%) • Salmonella typhi (killed or live attenuated) (80%) • Mycobacterium tuberculosis (BCG) (0-70%)Monday, September 10, 2012
  • VACCINE- PREVENTABLE DISEASESMonday, September 10, 2012
  • VACCINES FOR YOU Following primary series of immunizations, young adults should have a further dose of TD at age 15-16 years Health care workers need evidence of two measles vaccinations or blood test conforming immunity Meningitis vaccination recommended for students & staff in residential colleges of the university or in any group accommodationMonday, September 10, 2012
  • VACCINES FOR YOU Following primary series of immunizations, young adults should have a further dose of TD at age 15-16 years Health care workers need evidence of two measles vaccinations or blood test conforming immunity Meningitis vaccination recommended for students & staff in residential colleges of the university or in any group accommodationMonday, September 10, 2012
  • VACCINES FOR YOU Following primary series of immunizations, young adults should have a further dose of TD at age Basic Vaccinations: 15-16 years Tetanus Diphtheria Health care workers need evidence of Polio Measles two measles vaccinations or blood Mumps test conforming immunity Rubella Hepatitis B Meningitis vaccination recommended Meningitis for students & staff in residential colleges of the university or in any group accommodationMonday, September 10, 2012
  • VACCINES FOR YOU • RUBELLA • Lyophilized preparation of highly attenuated Wistar RA 27/3 strain rubella virus propagated in human diploid cells • 0.5ml single dose , SC • Side effects: rashes, malaise, temperature elevation, cough, coryza, headache, transient arthralgia & arthritis with or w/o joint effusionMonday, September 10, 2012
  • VACCINES FOR YOU • RUBELLA • Lyophilized preparation of highly attenuated Wistar RA 27/3 strain rubella virus propagated in human diploid cells • 0.5ml single dose , SC • Side effects: rashes, malaise, temperature elevation, cough, coryza, headache, transient arthralgia & arthritis with or w/o joint effusionMonday, September 10, 2012
  • VACCINES FOR YOU • RUBELLA Basic Vaccinations: • Lyophilized preparation of highly Tetanus attenuated Wistar RA 27/3 strain Diphtheria rubella virus propagated in human Polio diploid cells Measles Mumps • 0.5ml single dose , SC Rubella Hepatitis B Meningitis • Side effects: rashes, malaise, temperature elevation, cough, coryza, headache, transient arthralgia & arthritis with or w/o joint effusionMonday, September 10, 2012
  • VACCINES FOR YOU • MMR • Live attenuated measles (Edmonston –Zagreb strain), mumps (Rubini strain), rubella (Wistar RA 27/3 strain) viruses. • Strains propagated on human diploid cells, no antibiotics • Vaccinate infants only after 15 months = Repeat immunization after 15 years for infants immunized before age 12 monthsMonday, September 10, 2012
  • VACCINES FOR YOU • MMR • Live attenuated measles (Edmonston –Zagreb strain), mumps (Rubini strain), rubella (Wistar RA 27/3 strain) viruses. • Strains propagated on human diploid cells, no antibiotics • Vaccinate infants only after 15 months = Repeat immunization after 15 years for infants immunized before age 12 monthsMonday, September 10, 2012
  • VACCINES FOR YOU • MMR • Live attenuated measles (Edmonston Basic Vaccinations: –Zagreb strain), mumps (Rubini Tetanus strain), rubella (Wistar RA 27/3 Diphtheria strain) viruses. Polio Measles • Strains propagated on human diploid Mumps cells, no antibiotics Rubella Hepatitis B • Vaccinate infants only after 15 Meningitis months = Repeat immunization after 15 years for infants immunized before age 12 monthsMonday, September 10, 2012
  • VACCINES FOR YOU • CHICKEN POX (VARICELLA) • Live attenuated varicella vaccine • For healthy infants from age 9 months onward, healthy subjects , high risk patients & healthy close contacts, patients with acute leukemia, under immunosuppressive treatment, patients with planned organ transplantation, patients with chronic diseases & healthy close contacts • Dose recommended: 2 doses 4-6 weeks apart for adultsMonday, September 10, 2012
  • VACCINES FOR YOU • CHICKEN POX (VARICELLA) • Live attenuated varicella vaccine • For healthy infants from age 9 months onward, healthy subjects , high risk patients & healthy close contacts, patients with acute leukemia, under immunosuppressive treatment, patients with planned organ transplantation, patients with chronic diseases & healthy close contacts • Dose recommended: 2 doses 4-6 weeks apart for adultsMonday, September 10, 2012
  • VACCINES FOR YOU • CHICKEN POX (VARICELLA) Additional • Live attenuated varicella vaccine Vaccinations: • For healthy infants from age 9 months Chicken pox onward, healthy subjects , high risk Hepatitis A patients & healthy close contacts, patients Influenza Pertussis with acute leukemia, under immunosuppressive treatment, patients Salmonella typhi with planned organ transplantation, Cholera patients with chronic diseases & healthy Pneumonia close contacts • Dose recommended: 2 doses 4-6 weeks apart for adultsMonday, September 10, 2012
  • VACCINES FOR YOU • HEPATITIS A • Inactivated hepatitis A virus for individuals > 15 years of age in pre-filled syringe • Virosomal hepatitis A virus antigen (RG-SB strain) for adults & children over 12 months • Basic immunization : Intramuscular injection • Booster immunization: 6, 12 or 18 months after first immunization • Simultaneous active & passive immunization at another siteMonday, September 10, 2012
  • VACCINES FOR YOU • HEPATITIS A • Inactivated hepatitis A virus for individuals > 15 years of age in pre-filled syringe • Virosomal hepatitis A virus antigen (RG-SB strain) for adults & children over 12 months • Basic immunization : Intramuscular injection • Booster immunization: 6, 12 or 18 months after first immunization • Simultaneous active & passive immunization at another siteMonday, September 10, 2012
  • VACCINES FOR YOU • HEPATITIS A Additional • Inactivated hepatitis A virus for individuals > Vaccinations: 15 years of age in pre-filled syringe Chicken pox • Virosomal hepatitis A virus antigen (RG-SB Hepatitis A strain) for adults & children over 12 Influenza months Pertussis • Basic immunization : Intramuscular Salmonella typhi injection Cholera • Booster immunization: 6, 12 or 18 months Pneumonia after first immunization • Simultaneous active & passive immunization at another siteMonday, September 10, 2012
  • VACCINES FOR YOU • INFLUENZA • Highly purified surface antigens of influenza viruses types A & B constituted strains annually recommended by WHO • Polyvalent whole virus vaccine against strains of groups A & B • Purified split inactivated influenza virusMonday, September 10, 2012
  • VACCINES FOR YOU • INFLUENZA • Highly purified surface antigens of influenza viruses types A & B constituted strains annually recommended by WHO • Polyvalent whole virus vaccine against strains of groups A & B • Purified split inactivated influenza virusMonday, September 10, 2012
  • VACCINES FOR YOU • INFLUENZA Additional Vaccinations: • Highly purified surface antigens Chicken pox of influenza viruses types A & Hepatitis A B constituted strains Influenza annually recommended by Pertussis WHO Salmonella typhi • Polyvalent whole virus vaccine Cholera Pneumonia against strains of groups A & B • Purified split inactivated influenza virusMonday, September 10, 2012
  • VACCINES FOR YOU • CHOLERA • Orochol E Berna contains not < 2x109 viable organisms of the attenuated strain Vibrio CVD 103-HgR in a lyophilized form • For ORAL active immunization of adults & children >2 years against cholera • Dosage: Resuspend contents of both chambers at the same time in cold or lukewarm water, mix carefully for 5-10 seconds & drink immediately. • Do not resuspend in milk, juice or carbonated beverages.Monday, September 10, 2012
  • VACCINES FOR YOU • CHOLERA • Orochol E Berna contains not < 2x109 viable organisms of the attenuated strain Vibrio CVD 103-HgR in a lyophilized form • For ORAL active immunization of adults & children >2 years against cholera • Dosage: Resuspend contents of both chambers at the same time in cold or lukewarm water, mix carefully for 5-10 seconds & drink immediately. • Do not resuspend in milk, juice or carbonated beverages.Monday, September 10, 2012
  • VACCINES FOR YOU • CHOLERA • Orochol E Berna contains not < 2x109 viable organisms of the attenuated strain Vibrio CVD 103-HgR in a lyophilized form Additional Vaccinations: • For ORAL active immunization of adults & Chicken pox children >2 years against cholera Hepatitis A Influenza Pertussis • Dosage: Resuspend contents of both chambers at the same time in cold or Salmonella typhi lukewarm water, mix carefully for 5-10 Cholera seconds & drink immediately. Pneumonia • Do not resuspend in milk, juice or carbonated beverages.Monday, September 10, 2012
  • VACCINES FOR YOU • TYPHOID • Vi capsular polysaccharide typhoid vaccine: Single IM or SC injection • Oral Vaccine preparation > 109 viable organisms of attenuated Salmonella typhi strain Ty21a Berna; has cross immunity against S. paratyphi BMonday, September 10, 2012
  • VACCINES FOR YOU • TYPHOID • Vi capsular polysaccharide typhoid vaccine: Single IM or SC injection • Oral Vaccine preparation > 109 viable organisms of attenuated Salmonella typhi strain Ty21a Berna; has cross immunity against S. paratyphi BMonday, September 10, 2012
  • VACCINES FOR YOU • TYPHOID Additional • Vi capsular polysaccharide Vaccinations: typhoid vaccine: Single IM or SC Chicken pox injection Hepatitis A Influenza Pertussis • Oral Vaccine preparation > 109 Salmonella typhi viable organisms of attenuated Cholera Pneumonia Salmonella typhi strain Ty21a Berna; has cross immunity against S. paratyphi BMonday, September 10, 2012
  • VACCINES FOR YOU • PNEUMOCOCCUS • Purified polysaccharide of Streptococcus pneumoniae • 25ug each of 23 serotypes: 1,2,3,4,5,6B,7F,8,9V,10A, 11A, 12F, 14, 15B, 17F, 18C, 19A, 19F, 20, 22F, 23F, 33F • Indications: Prevention of pneumococcal infections, particularly those of respiratory origin for ages over 2 years who are at risk of serious pneumococcal infectionMonday, September 10, 2012
  • VACCINES FOR YOU • PNEUMOCOCCUS • Purified polysaccharide of Streptococcus pneumoniae • 25ug each of 23 serotypes: 1,2,3,4,5,6B,7F,8,9V,10A, 11A, 12F, 14, 15B, 17F, 18C, 19A, 19F, 20, 22F, 23F, 33F • Indications: Prevention of pneumococcal infections, particularly those of respiratory origin for ages over 2 years who are at risk of serious pneumococcal infectionMonday, September 10, 2012
  • VACCINES FOR YOU • PNEUMOCOCCUS • Purified polysaccharide of Streptococcus pneumoniae Additional Vaccinations: Chicken pox • 25ug each of 23 serotypes: Hepatitis A 1,2,3,4,5,6B,7F,8,9V,10A, 11A, 12F, 14, Influenza 15B, 17F, 18C, 19A, 19F, 20, 22F, 23F, Pertussis 33F Salmonella typhi • Indications: Prevention of pneumococcal Cholera infections, particularly those of Pneumonia respiratory origin for ages over 2 years who are at risk of serious pneumococcal infectionMonday, September 10, 2012
  • VACCINES FOR YOU • PNEUMOCOCCUS • Dosage: Initial Injections: Single IM or SC injection confers protection against the 23 types of pneumococci in the vaccine • Revaccination should not be carried out for at least 5 years as severe local reactions (Arthus phenomena type) were observed if time between 2 injections is too shortMonday, September 10, 2012
  • VACCINES FOR YOU • PNEUMOCOCCUS • Dosage: Initial Injections: Single IM or SC injection confers protection against the 23 types of pneumococci in the vaccine • Revaccination should not be carried out for at least 5 years as severe local reactions (Arthus phenomena type) were observed if time between 2 injections is too shortMonday, September 10, 2012
  • VACCINES FOR YOU • PNEUMOCOCCUS • Dosage: Initial Injections: Single Additional IM or SC injection confers Vaccinations: Chicken pox protection against the 23 types of Hepatitis A pneumococci in the vaccine Influenza Pertussis • Revaccination should not be Salmonella typhi carried out for at least 5 years as Cholera severe local reactions (Arthus Pneumonia phenomena type) were observed if time between 2 injections is too shortMonday, September 10, 2012
  • ISSUES TO CONSIDER • Which vaccine to administer to whom? • When to vaccinate: months before exposure? • How many doses? Cost? • Interval between doses in case of multi-dose regimen for primary immunization • When to expect protection? Immunologic response vs protection? • Possibility of adverse/side effectsMonday, September 10, 2012
  • EXPANDED IMMUNIZATION PROGRAMSMonday, September 10, 2012
  • The 7 EPI Diseases Measles Diphtheria Poliomyelitis Hepatitis B Pertussis Tetanus TuberculosisMonday, September 10, 2012
  • 1976 Official launching of EPI in the Philippines 1977 BCG & DPT2 for infants in priority areas BCG & DPT2 expanded nationwide 1979 OPV3 in selected areas reporting outbreaks TT2 for pregnant women at 5 months gestation 1980 OPV3 & TT2 given nationwide 1982 Measles vaccine for 35% of the population 1983 Measles vaccination given nationwide 1984 DPT3 added; TT given anytime during pregnancy 1985 Comprehensive Program ReviewMonday, September 10, 2012
  • 1986 Universal Child Immunization Goal by 1990-UN Start of Rotary International mass campaigns –Polio 1987 Plus TT3, TT4, TT5 added; Wednesday adopted as 1989 Immunization Day National Plan of Action for Polio Eradication 1991 launched; Polio Eradication Unit 1992 Hepatitis B immunization -40% of infants 1993 1st National Immunization Days Neonatal Tetanus Elimination-Action Plan Measles Control – National Plan of ActionMonday, September 10, 2012
  • Bacteria Disease Antigen Efficacy Streptococcus Capsular Pneumonia 85% pneumoniae polysaccharide Neisseria Capsular meningitis 85% meningitidis polysaccharide Hemophilus Capsular meningitis 85% influenzae polysaccharide Corynebacterium Diphtheriae Toxoid >90% diphtheriae Clostridium tetani tetanus Toxoid >90% Bordetella Whooping cough Killed organism >90% pertussis Mycobacterium Tuberculosis Live attenuated 0 – 70 % bovis (BCG) Killed/Live Salmonella typhi Typhoid fever 80% attenuatedMonday, September 10, 2012
  • Vaccine Reason for Immunization Given at the earliest possible age protects BCG against the possibility of infection from family members Early start with DPT reduces chance of DPT severe pertussis Protection against polio increased the OPV earlier the OPV is given Early start of HBV vaccination reduces Hepatitis B chance of being infected and becoming a carrier At least 80% of measles can be prevented Measles by immunization at this ageMonday, September 10, 2012
  • Diseases to be immunized Age against Birth Tuberculosis 6 weeks DPT; Polio; HBV 10 weeks DPT; Polio; HBV 14 weeks DPT; Polio; HBV 9 months MeaslesMonday, September 10, 2012
  • Tetanus toxoid Immunization Schedule for Women Percent Vaccine Minimum age/interval Duration of protection protected As early as possible during TT1 pregnancy * Infants born to the mother protected from TT2 At least 4 weeks later 80% neonatal tetanus; 3 years protection to mother * Infants protected; TT3 At least 6 months later 95% 5 years protection to mother * Infants protected; TT4 At least one year later 99% 10 years protection to mother Lifetime protection for mother; * all infants born TT5 At least one year later 99% to that mother will be protectedMonday, September 10, 2012
  • Philippine Immunization Flagship Programs  Poliomyelitis Eradication  Measles Elimination  Maternal and Neonatal Tetanus Elimination  Control of Hepatitis BMonday, September 10, 2012
  • TAKE-HOME QUIZ • Next to Smallpox, why do you think that polio and measles are the next diseases targeted for global elimination? • Differentiate the 4 Immunization Flagship Programs of the country in terms of: • target group, vaccine to use and route, criteria for successMonday, September 10, 2012
  • IMMUNOLOGIC TOLERANCE AND AUTOIMMUNITYMonday, September 10, 2012
  • RECALL... • Immunologic tolerance: unresponsiveness to self-antigens • WHAT WE WILL LEARN: • How does the immune system maintain its unresponsiveness to self-antigens • What are the factors that may contribute to the development of autoimmunity?Monday, September 10, 2012
  • ON ENCOUNTER WITH LYMPHOCYTES...Monday, September 10, 2012
  • ON ENCOUNTER WITH LYMPHOCYTES... The choice among lymphocyte activation, tolerance, and ignorance is determined by the nature of the antigen-specific lymphocytes and by the nature of the antigen and how it is displayed to the immune systemMonday, September 10, 2012
  • IMPORTANCE OF IMMUNOLOGIC TOLERANCE • the knowledge that self-antigens normally induce tolerance • if we learn how to induce tolerance in lymphocytes specific for a particular antigen, we may be able to use this knowledge to prevent or control unwanted immune reactions • APPLICATIONS: • treat allergic and autoimmune diseases • prevent the rejection of organ transplants • gene therapy • prevent immune responses against the products of newly expressed genes or vectorsMonday, September 10, 2012
  • CENTRAL & PERIPHERAL • central tolerance: induced when developing lymphocytes encounter these antigens in the generative lymphoid organs (bone marrow and thymus) • peripheral tolerance: induces when mature lymphocytes encounter self antigens in peripheral tissuesMonday, September 10, 2012
  • AUTOIMMUNITY • Principal factors in the development of autoimmunity: • inheritance of susceptibility genes • may contribute to failure of self-tolerance • environmental triggers • e.g. infections = may activate self-reactive lymphocytesMonday, September 10, 2012
  • CENTRAL T-LYMPHOCYTE TOLERANCE • Negative selection: principal mechanism of central tolerance • Strong recognition of self antigens immature T-cells in the thymus may lead to death of the cells (negative selection or deletion) • Self-antigen recognition in the thymus may also lead to the development of regulatory T- cells that enter peripheral tissuesMonday, September 10, 2012
  • PERIPHERAL T- LYMPHOCYTES & ANERGY • Peripheral tolerance: • induced when mature T cells recognize self antigens in peripheral tissues, leading to functional inactivation (anergy) or death • induced when the self-reactive lymphocytes are suppressed by regulatory T cellsMonday, September 10, 2012
  • ANERGY • ANERGY: functional inactivation of T-lymphocytes that occurs when these ceIls recognize antigens without adequate levels of the costimulators (second signals) that are needed for full T-cell activationMonday, September 10, 2012
  • ACTIVATION-INDUCED CELL DEATH : DELETIONMonday, September 10, 2012
  • IMMUNE SUPPRESSIONMonday, September 10, 2012
  • Characteristics of self and foreign (e.g. microbial) protein antigens that determine why the self antigens induce tolerance and microbial antigens stimulate T-cell mediated immune responsesMonday, September 10, 2012
  • CENTRAL TOLERANCE IN B-CELLSMonday, September 10, 2012
  • PERIPHERAL TOLERANCE IN B-CELLSMonday, September 10, 2012
  • AUTOIMMUNE DISEASES & MHC MOLECULESMonday, September 10, 2012
  • Monday, September 10, 2012
  • MOLECULAR MIMICRY & AUTOIMMUNITYMonday, September 10, 2012
  • SUMMARY • Immunologic tolerance: specific unresponsiveness to an antigen induced by exposure of lymphocytes to that antigen • All individuals are tolerant of (unresponsive to) their own (self)antigens • Tolerance against antigens may be induced by administering that antigen in particular ways = strategy may be useful for treating immunologic disease and for preventing the rejection of transplantsMonday, September 10, 2012
  • SUMMARY • Autoimmune diseases: result from a failure of self- tolerance • Multiple factors contribute to autoimmunity: • immunologic abnormalities • susceptibility genes • infectionsMonday, September 10, 2012
  • SUMMARY • Central tolerance: induced by the death of immature lymphocytes that encounter antigens in the generative lymphoid organs while Peripheral tolerance: results from the recognition of antigens by mature Iymphocytes in peripheral tissues • T-CELLS: • Central tolerance (negative selection): result of high- affinity recognition of antigens in the thymus, which tend to be widely disseminated self-antigens AND may eliminate the potentially most dangerous T-cells, which express high- affinity receptors for disseminated self-antigensMonday, September 10, 2012
  • SUMMARY • T-CELLS: • Peripheral tolerance: induced by multiple mechanisms • Anergy (functional inactivation) results from the recognition of antigens without costimulators (second signals) or when T cells use inhibitory receptors to recognize costimulators • Deletion (death by apoptosis) occurs when T cells repeatedly encounter self antigens • Some self-reactive T cells suppress potentially pathogenic T-cellsMonday, September 10, 2012
  • SUMMARY •B-CELLS • central tolerance: induced when immature cells recognize self antigens in the bone marrow • peripheral tolerance: (by anergy) induced when mature B cells recognize self antigens without T cell helpMonday, September 10, 2012
  • SUMMARY • AUTOIMMUNITY • Many genes contribute to the development of autoimmunity • The strongest associations are between HLA genes and various T cell-mediated autoimmune diseases • Infections predispose to autoimmunity, by causing inflammation and inducing the aberrant expression of costimulators or because of cross-reactions between microbial and self antigensMonday, September 10, 2012
  • vanderbilt.edu IMMUNE RESPONSE AGAINST TUMORS/TRANSPLANTS Reference; Immunology by AbbasMonday, September 10, 2012
  • CALENDAR FOR BIO 151 DATE ACTIVITY September 11 TUMORS, TRANSPLANTS & SERODIAGNOSIS September 18 EXAMINATION 2 (VACCINES TO SEROLOGICAL DIAGNOSTICS) September 25- October 2 IMMUNITY TO MICROBIAL INFECTIONS: Pneumonia a.Fungi b.Virus c.Bacteria IMMUNITY TO MICROBIAL INFECTIONS: Schistosomiasis October 9 EXAMINATION 3: Take-Home (24 hours) Questions available October 9, 2012 at 12 noon...Submission should be October 10, 2012 at 12 noon (at DB, received and stamped by DB staff) *** Printed and well-citedMonday, September 10, 2012
  • WHAT ARE TUMORS? webmd.com • Simply means a mass of cells • Can be either benign or malignant • Benign tumors: not a threat to life or long- term health; made up of related but different cells • Malignant tumors: threat to life or long- term health; clonal or all identical to the cell of originMonday, September 10, 2012
  • IMMUNE RESPONSE TO TUMORS • T cells generally mount effective surveillance against tumors associated with oncogenic viruses or UV induction (these are strongly immunogenic) • More weakly immunogenic tumors are not controlled by joseph-birch.livejournal.com T cell surveillance, although sometimes low grade responses are evokedMonday, September 10, 2012
  • IMMUNE RESPONSE TO TUMORS • NK cells play a role in containing tumor growth and metastases; • The importance of the immune system in preventing tumor growth can be seen in immunocompromised patients - AIDS patients have a much higher chance of developing many types of cancerMonday, September 10, 2012
  • WHAT IS CANCER? • diseases in which there is loss of regulation of the proliferative process • characterized by excessive, uncontrolled growth of abnormal cells, which invade and destroy other tissues nursingcrib.com • hyperproliferation of cells that have violated the basic rules of social cell behavior (i.e, lost the ability to be controlled by normal cell signals)Monday, September 10, 2012
  • CANCER • Control is important to ensure that cells only divide when needed (organs and tissues should maintain their correct shapes and sizes) • Should this system fail, several backup safety mechanisms prevent topnews.in the cell from dividing uncontrollablyMonday, September 10, 2012
  • DEFENSE AGAINST UNCONTROLLED PROLIFERATION • A cell can initiate cell death (suicide) when a defect is detected through a process called apoptosis • A cell can repair the error and become a normal cell genengnews.com • Immune system (e.g., natural killer cells can detect abnormal cells and kill them)Monday, September 10, 2012
  • WHAT ARE ORGANS TRANSPLANTS? • surgical operation in which a failing or damaged organ in the human body is removed and replaced with a functioning one • donated organ may be from a deceased donor, a living donor, or an animal • NOTE: In some cases an artificial organ is usedMonday, September 10, 2012
  • WHAT ARE ORGANS TRANSPLANTS? • Cadaveric organ donation involves removing organs from a recently deceased donor • Living organ donation involves the donation of one of a paired organ (such as kidneys) or a portion of an organ (such as a lobe of the liver or lung) • The donors organ system is still able to function after the donation • Living donors are often related to the patient, but that is not always the caseMonday, September 10, 2012
  • SO WHAT’S IT FOR IMMUNOLOGY? • CANCER: enhancing immunity against the tumors holds much promise for treatment •TRANSPLANTS: immune responses against transplants are a barrier for a successful transplantationMonday, September 10, 2012
  • WHAT WE WILL LEARN... • What are the antigens in tumors and tissue transplants that are recognized as foreign by the immune system? • How does the immune system recognize and react to tumors and transplants? • How can the immune responses to tumors and grafts be manipulated to enhance tumor rejection and inhibit graft rejection?Monday, September 10, 2012
  • IMMUNE SURVEILLANCE • 1950s: a physiologic function of the adaptive immune system is to prevent the outgrowth of transformed cells or to destroy these cells before they become harmful tumors london-research-institute.org.uk aihealthsolutions.caMonday, September 10, 2012
  • EVIDENCES SUPPORTING THE CONCEPT OF IMMUNO SURVEILLANCEMonday, September 10, 2012
  • TUMOR ANTIGENS • Malignant tumors express various types of molecules that may be recognized by the immune system as foreign antigensMonday, September 10, 2012
  • IMMUNE MECHANISM FOR TUMOR REJECTION • The principal immune mechanism of tumor eradication is killing of tumor cells by cytolytic T-lymphocytes (CTLs) specific for tumor antigens • endogenously synthesized cytosolic proteins displayed as Class I MHC-associated peptides • SO: recognized by class I MHC-restricted CD8+ CTLs (kills the cell producing the antigens)Monday, September 10, 2012
  • IMMUNE MECHANISM FOR TUMOR REJECTION • CTLs responses against tumor cells are often induced by recognition of tumor antigens on host-antigen presenting cells (APCs) which ingest tumor cells or their antigens and present the antigens to T-cellsMonday, September 10, 2012
  • IMMUNE MECHANISM FOR TUMOR REJECTION • anti-tumor CD4+ T- cell responses and antibodies have been detected in patients (but may not be protective) • activated macrophages and NK cells are capable of killing tumor cells in vitro (unclear protection)Monday, September 10, 2012
  • EVASION OF IMMUNE RESPONSE BY TUMORS • Immune response often fail to check tumor growth because these responses are ineffective or because tumors evolve to evade immune attack • Immune responses against tumors may be weak because many tumor antigens are weakly immunogenic, perhaps because they differ only slightly from self antigensMonday, September 10, 2012
  • EVASION OF IMMUNE RESPONSE BY TUMORS • Growing tumors also develop mechanisms for evading immune responses • stop expressing the antigens that are targets of immune attack (antigen loss variants) = continue to grow and spread • stop expressing class I MHC molecules = cannot display antigens to CD8+ T-cells (cross- priming) • may produce molecules, e.g. transforming growth factor-beta, that suppress immuneMonday, September 10, 2012 responses
  • IMMUNOTHERAPY FOR TUMORS • MAIN GOAL: • provide antitumor effectors (antibodies and T-cells) to patients • actively immunize patients against their tumors • stimulate the patients’ own antitumor immune responseMonday, September 10, 2012
  • IMMUNOTHERAPY VS CHEMOTHERAPY • Chemotherapy/ Irradiation: may have devastating effects on normal non-tumor tissues • Immunotherapy: highly- specific for the tumor health.umn.eduMonday, September 10, 2012
  • STRATEGY 1: PASSIVE IMMUNITY • immune effectors are injected into cancer patients • e.g. monoclonal antibodies to tumor antigens + potent toxins • HOW: antibodies bind to tumor antigens and activate host effector mechanisms (e.g. phagocytes or complement system) • HOW: toxins delivered to tumor cellsMonday, September 10, 2012
  • EXAMPLE: BREAST CANCER IMMUNOTHERAPY • ALL information from: Critical Reviews in Oncology and Hematology, October 2010 (Emde et al. Therapeutic strategies and mechanisms of tumorigenesis of HER2-overexpressing breast cancer) • HER2 (tyrosine kinase): overexpressed in approximately 25% of breast cancers; acts as a signal amplifier for its siblings = transmembrane receptors that collectively bind with 11 distinct growth factors of the EGF family • THUS: overexpression of HER2 confers aggressive invasive growth in preclinical models and in patients • “Specific therapies targeting HER2 include monoclonal antibodies, antibody–drug conjugates, small molecule tyrosine kinase inhibitors, as well as heat shock protein and sheddase inhibitors. Two of these drugs have shown impressive – yet mostly transient – efficacy in patients with HER2 overexpressing breast cancer.”Monday, September 10, 2012
  • Signal transduction pathways instigated by HER2, co-receptors and EGF-like growth factors (epidermal) and NRGs (neuregulins)Monday, September 10, 2012
  • Clinically approved and experimental therapeutic strategies targeting ErbB-2/ HER2 in carcinomas: a. Trastuzumab: humanized monoclonal antibody directed against the extracellular domain of HER2; recruits immune effector mechanisms and can induce apoptosis, block angiogenesis and inhibit tumor cell proliferation. b. Pertuzumab: prevent heterodimerization of HER2 with other family members RECENTS: a. Lapatinib: tyrosine kinase inhibitors; reversible inhibitor b. Neratinib: irreversible inhibitor; variably inhibit a broad range of tyrosine kinases c. 17-AAG: block the ATP/ADP binding pocket of HSP90 and target HER2 for proteasomal degradation (Note; HSP90: molecular chaperone required for proper folding of protein kinases like HER2Monday, September 10, 2012
  • STRATEGIES FOR ENHANCING ANTITUMOR IMMUNE RESPONSE • 1. Tumor- antigen pulsed DC: mimic normal pathway of cross- presentation to induce specific T-cell responseMonday, September 10, 2012
  • STRATEGIES FOR ENHANCING ANTITUMOR IMMUNE RESPONSE • 2. DNA or transfected DC: the host produce the tumor antigen thus inducing specific T- cell responseMonday, September 10, 2012
  • STRATEGIES FOR ENHANCING ANTITUMOR IMMUNE RESPONSE • 3. tumor cell expressing costimulators (e.g. B7) or cytokines (e.g. IL-2): cytokines or costimulators to stimulate the generation of own tumor-specific immune responseMonday, September 10, 2012
  • IMMUNE RESPONSE AGAINST TRANSPLANTSMonday, September 10, 2012
  • IMMUNE RESPONSE AGAINST TRANSPLANTS • transplants exchanged wenliang.myweb.uga.edu between animals of the same and other inbred are accepted • grafts among different strains are rejected • GENES for graft rejection: products are expressed in ALL tissuesMonday, September 10, 2012
  • IMMUNE RESPONSE AGAINST TRANSPLANTS • Allogeneic (allografts) and Xenogeneic (xenografts) grafts are always rejected • Alloantigens (alloreactive antibodies and T-cells) • Xenoantigens (xenoreactive antobodies and T-cells) • CLINICAL Scenario: mostly allogeneicMonday, September 10, 2012
  • Transplantation Antigens • The antigens of allografts that serve as the principal targets of rejection are proteins encoded in the MHC • Human MHC = human leukocyte antigen or HLAMonday, September 10, 2012
  • INDUCTION OF IMMUNE RESPONSES AGAINST TRANSPLANTS • T-cells may recognize allogeneic MHC molecules in the graft by professional APCs, or graft alloantigens may be processed and presented by hosts’ professional APCsMonday, September 10, 2012
  • INDUCTION OF IMMUNE RESPONSES AGAINST TRANSPLANTS • T-cells may recognize allogeneic MHC molecules if the graft does not contain professional APCs, how does it stimulate T cells? in the graft by professional APCs, or graft alloantigens may be processed and presented by hosts’ professional APCsMonday, September 10, 2012
  • INDUCTION OF IMMUNE RESPONSES AGAINST TRANSPLANTS • T-cells may recognize allogeneic MHC molecules if the graft does not contain professional APCs, how does it stimulate T cells? in the graft by professional APCs, or graft alloantigens may be processed and presented by hosts’ graft cells are ingested by professional APCs in the recipient and professional the donor alloantigens are processed and presented by the self MHC molecules on recipient APCs APCsMonday, September 10, 2012
  • MIXED LYMPHOCYTE REACTION (MLR) wenliang.myweb.uga.edu • an in vitro model of T-cell recognition in alloantigensMonday, September 10, 2012
  • MIXED LYMPHOCYTE REACTION (MLR) • T-cells from one individual are cultured with leukocytes of another individual and the responses of the T-cells are assayed • magnitude of response is proportional to the extent of MHC differences between these individuals • rough predictor of the outcomes of grafts exchanged between these individualsMonday, September 10, 2012
  • IMMUNE MECHANISM OF GRAFT REJECTION • occurs within minutes of transplantation; characterized by thrombosis of graft vessels and ischemic necrosis of the graft • mediated by circulating antibodies specific for antigens on graft endothelial cellsMonday, September 10, 2012
  • IMMUNE MECHANISM OF GRAFT REJECTION not common BUT major barrier to xenotransplantation • occurs within minutes of transplantation; characterized by thrombosis of graft vessels and ischemic necrosis of the graft • mediated by circulating antibodies specific for antigens on graft endothelial cellsMonday, September 10, 2012
  • IMMUNE MECHANISM OF GRAFT REJECTION • occurs within days or weeks after transplantation • principal cause of early graft failure • mediated mainly y T-cells which react against alloantigens in the graft or may react agains cells in graft vessels leading to vascular damageMonday, September 10, 2012
  • IMMUNE MECHANISM OF GRAFT REJECTION antibodies may also play a role • occurs within days or weeks after transplantation • principal cause of early graft failure • mediated mainly y T-cells which react against alloantigens in the graft or may react agains cells in graft vessels leading to vascular damageMonday, September 10, 2012
  • IMMUNE MECHANISM OF GRAFT REJECTION • occurs over months or years that leads to progressive loss of graft function • manifested as fibrosis or narrowing of vessels (arteriosclerosis) • T-cells secrete cytokines which stimulate proliferation of fibroblasts and vascular smooth muscle cells in the graftMonday, September 10, 2012
  • PREVENTION & TREATMENT OF GRAFT REJECTION e-steroid.comMonday, September 10, 2012
  • BONE MARROW TRANSPLANTS • elicit strong rejection reactions: must perform cross- matches • carry the risk of graft-versus- host disease: If mature allogeneic T cells are transplanted with the marrow cells, these mature T cells can attack the recipients tissues • lead to temporary immunodeficiency in recipientsMonday, September 10, 2012
  • QUESTIONS?Monday, September 10, 2012
  • elitechgroup.com SERODIAGNOSIS Reference; Jawetz Medical Microbiology (http://accessmedicine.com/resourceTOC.aspx?resourceID=6)Monday, September 10, 2012
  • ehrs.upenn.edu BIOSAFETY 2008.igem.org busytrade.comMonday, September 10, 2012
  • OVERVIEW OF DIAGNOSTICS • 3 CATEGORIES: • Direct: clinical specimen is examined directly for the presence of virus particles, virus antigen or viral nucleic acids • Indirect: the specimen into cell culture, eggs or animals in an attempt to grow the virus • Serology: detection of rising titres of antibody between acute and convalescent stages of infection, or the detection of IgMonday, September 10, 2012
  • ANTIGEN DETECTION: DIRECT • Applications: • immunofluorescence testing of nasopharyngeal aspirates for respiratory viruses e.g.. RSV, flu A, flu B, and adenoviruses • detection of rotavirus antigen in fecesrotavirus, adenovirus, astroviruses, Norwalk-like viruses • the pp65 CMV antigenaemia test • the detection of HSV and VZV in skin scrapingMonday, September 10, 2012
  • ANTIGEN DETECTION: DIRECT geneaid.com wclassproducts.com norgenbiotek.comMonday, September 10, 2012
  • ANTIGEN DETECTION: INDIRECTMonday, September 10, 2012
  • ANTIGEN DETECTION: INDIRECT (enhanced)Monday, September 10, 2012
  • ANTIBODY DETECTION: SEROLOGY http://virology-online.com usada.orgMonday, September 10, 2012
  • COMMON TESTS • Classical Techniques: • Newer Techniques: • Complement fixation tests • Radioimmunoassay (RIA) (CFT) • Enzyme linked • Hemagglutination inhibition immunosorbent assay (EIA) tests (HAI) • Particle agglutination • Immunofluorescence techniques (IF) • Western Blot (WB) • Neutralization tests • Recombinant immunoblot assay (RIBA) • Single Radial HemolysisMonday, September 10, 2012
  • DIAGNOSING A PRIMARY (ACUTE) INFECTION • A significant rise in titre of IgG/total antibody between acute and convalescent sera • CFT and HAI, it is normally taken as a four-fold or greater increase in titre • diagnosis is usually retrospective because by the time the convalescent serum is taken, the patient had probably recoveredMonday, September 10, 2012
  • DIAGNOSING A PRIMARY (ACUTE) INFECTION • Presence of IgM • EIA, RIA, and IF may be used (rapid) • BUT: • interference by rheumatoid factor, • re-infection by the virus • unexplained persistence of IgM years after the primary infectionMonday, September 10, 2012
  • DIAGNOSING A PRIMARY (ACUTE) INFECTION • Seroconversion • changing from a previously antibody negative state to a positive state • e.g. seroconversion against HIV following a needle- stick injury, or against rubella following contact with a known caseMonday, September 10, 2012
  • DIAGNOSING A PRIMARY (ACUTE) INFECTION • A single high titre of IgG (or total antibody) • this is a very unreliable means of serological diagnosis since the cut-off is very difficult to defineMonday, September 10, 2012
  • DIAGNOSING A RE- INFECTION / RE-ACTIVATION • It is often very difficult to differentiate re-infection/re- activation from a primary infection • When important: rubella infection in the first trimester of pregnancy: primary infection is associated with a high risk of fetal damage whereas re-infection is not • RE-INFECTION: sharp large rise in antibody titres • RE-INFECTION/ RE-ACTIVATION: IgM is usually low or absentMonday, September 10, 2012
  • USE DEPENDS ON PATHOGEN • onset of clinical symptoms coincide with the development of antibodies: e.g. rubella and hepatitis A (IgM or rising titres of IgG in the serum of the patient would indicate active disease) • pathogen produce clinical disease before the appearance of antibodies: e.g. respiratory and diarrheal viruses (serological diagnosis would be retrospective and therefore will not be that useful) • pathogen produce clinical disease months or years after seroconversion: e.g. HIV and rabies (the mere presence of antibody is sufficient to make a definitive diagnosis)Monday, September 10, 2012
  • READING ASSIGNMENT: Q: WHAT DO YOU THINK ARE SOME PROBLEMS WITH SEROLOGYMonday, September 10, 2012
  • medicineworld.org THE SEROLOGICAL immunology.utoronto.ca DIAGNOSTICSMonday, September 10, 2012
  • COMPLEMENT FIXATION TEST http://virology-online.com path.cam.ac.uk dshs.state.tx.usMonday, September 10, 2012
  • HEMAGGLUTINATION/ HEMAGGLUTINATION INHIBITION TESTS (HAI) motifolio.com virology.wsMonday, September 10, 2012
  • HEMAGGLUTINATION/ HEMAGGLUTINATION INHIBITION TESTS (HAI) qmark2.leeds.ac.uk path.cam.ac.ukMonday, September 10, 2012
  • IMMUNOFLUORESCENCE TECHNIQUE (IF) • DIRECT: known labeled antibody interacts with unknown antigen • INDIRECT: known antigen is attached to a slide, unknown serum is added and then washed = An/Ab fixed onto slide and can be detected by antibody-specific reagentMonday, September 10, 2012
  • IMMUNOFLUORESCENCE TECHNIQUE (IF) Respiratory Syncytial Virus (RSV) (a) Indirect technique: polyclonal rabbit serum and a conjugated sheep anti-rabbit serum; both were extensively absorbed to remove unwanted antibodies (b) Direct technique: mixture of three monoclonal conjugated antibodies and Evans blue counterstain (which, paradoxically, appears red) = staining only those antigenic concentrations for which there are antibodies in the mixture CREDITS: (Both micrographs were taken by Joyce McQuillin personally, SCIENCE DIRECT)Monday, September 10, 2012
  • NEUTRALIZATION TESTS • potential of the serum of neutralizing the biological http://www.sanidadanimal.info activity of an antigen can also be known • assessment of the capability of a serum against bacterial toxins, bacteria or viruses biobest.co.uk • laborious, need cell cultures, sterile conditions, and usually require more timeMonday, September 10, 2012
  • NEUTRALIZATION TESTS Positive MDCC-MSB1 infected cells culture showing fluorescence at the point where CAV antigen was detected biobest.co.uk culture negative to CAMonday, September 10, 2012
  • SINGLE RADIAL HEMOLYSIS • E.g. Rubella antibody. • LEFT PLATE: Control Plate • RIGHT PLATE: well in the middle of the plate contains the 15 miu/ml control serum = clear zone of lysis surrounding the well on the test plate which is absent on the control plate • NOTE: Specimens which gives a zone of lysis equal or greater than the 15 miu/ml control well are regarded as positive for rubella antibody. = Should a zone of similar size is present on the control plate, then the result is not validMonday, September 10, 2012
  • RADIOIMMUNOASSAY (RIA) • used to quantitate antigens or haptens taht can be radioactively-labeled • based on competition for specific antibody between the labeled (known) and the unlabeled (unknown) concentration of the material • complex that form between An/Ab can then be separated and the amount of radioactivity measuredMonday, September 10, 2012
  • RADIOIMMUNOASSAY (RIA) • concentration of the unlabeled (unknown) antigen or hapten is determined by comparing the results with those obtained using several concentrations of a predetermined standard antigen • highly sensitive for: • assay of hormones or drugs from serum • measure amount of IgE with known allergenMonday, September 10, 2012
  • users.rcn.comMonday, September 10, 2012
  • RADIOALLERGOSORBENT ASSAY (RAST) lookfordiagnosis.com pennstatehershey.adam.comMonday, September 10, 2012
  • ENZYME IMMUNOASSAY (EIA) • depends on the conjugation of an enzyme to either an antigen or an antibody • enzyme is detected by assaying for enzyme activity with its substrate • no need for radioactive labels piercenet.comMonday, September 10, 2012
  • ELISA technologyinscience.blogspot.com • can detect the ANTIGEN- ANTIBODY reaction by the use of enzyme- linked antibodies • based  on the use of labeled antibodies (usually done with the enzyme peroxidase) so that the resulting conjugates have both immunological  and enzymatic activityMonday, September 10, 2012
  • ELISA • one of the components of the conjugate (antibody or antigen) is attached to the plate, so the antigen-antibody reaction can be easily measured when adding the reaction substrate • This substrate produces a colored reaction product when it comes into contact with the enzyme • The color change can be seen or quantified with a colorimeterMonday, September 10, 2012
  • ELISA: ANTIGEN DETECTION • Sandwich ELISA • plates are usually coated with an antibody (monoclonal or polyclonal antibody) against the unknown antigen • sample that needs to be tested is added to the wells, and if the antigen is present, the antigen-antibody reaction will take place • add another antibody linked to an enzyme. • when the reaction substrate is added it turns a colorMonday, September 10, 2012
  • ELISA: ANTIBODY DETECTION • INDIRECT ELISA: coating of the ELISA plate with the antigen against the specific antibodies that may be present in the serum • NEXT: addition of serum, incubation and washing;  addition of the conjugate, incubation and washing and finally, the addition of the substrate, stopping the reaction and reading the resultsMonday, September 10, 2012
  • ELISA: ANTIBODY DETECTION • COMPETITIVE ELISA: have an antibody (monoclonal of polyclonal) of a known antigen which has previously been bound to the plate • competitive because the serum is incubated with the antigen previous to its incubation with the antiserum bound to the plate = both compete for the antigen • NEXT: addition of the conjugate, incubation, washing, and finally, substrate addition and reading the resultsMonday, September 10, 2012
  • ELISA & HIV virology-online.com hivinfosource.orgMonday, September 10, 2012
  • DENGUE SEROLOGYMonday, September 10, 2012
  • QUESTIONS?Monday, September 10, 2012
  • END OF EXAM 2 COVERAGEMonday, September 10, 2012