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Allergies Overview
This document discusses different types of allergy testing for pets including intradermal testing and serology tests like ELISA, RAST, and western blot. It notes that serology offers advantages like being less invasive through a simple blood test and able to be done in one visit. Intradermal testing requires multiple visits and weaning off suppression medications. The document also covers suppression medications versus hyposensitization and their risks and success rates.
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Allergies Overview
- 1. SEROLOGY ALLERGY TESTINGSOTHER OPTION
- 2. OVERVIEW WHAT ISAN ALLERGY? SYMPTOMS SUPPRESSION VS HYPOSENSITIZATION WHEN TO ORDER TYPES OF TESTING HOT TOPICS IN ALLERGIES TODAY Q&A
- 3. WHAT IS ANALLERGY?
- 4. SYMPTOMS: SKIN ISSUESRESPIRATORY ISSUES DIGESTIVE PROBLEMS OCULAR SYMPTOMS
- 5. SKIN ISSUES ITCHINGREDNESS HOTSPOTS HAIR PULLING
- 6. RESPIRATORY ISSUES: COUGHINGSNEEZING WHEEZING REVERSE COUGH
- 7. DIGESTIVE ISSUES: VOMITINGDIARRHEA
- 8.
- 9. SUPPRESSION VS HYPOSENSITIZATIONSUPPRESSION: STEROIDS CYCLOSPORINE ANTIHISTAMINES HYPOSENSITIZATION: SMALL, CONTROLLED AMOUNTS OF OFFENDING ALLERGENS GIVEN SUBCUTANEOUSLY IN INCREASING DOSES AND FREQUENCY; PLANT DERIVED.
- 10. SUPPRESSION SIDE EFFECTSSTEROIDS DOGS: WEIGHT GAIN, HYPERACTIVITY, PANTING, DIARRHEA & DEPRESSION LIVER ADRENAL GLANDS PANCREAS DIABETES CATS: REQUIRE HIGHER DOSES TO ACHIEVE RESPONSE =COST LESS LIKELY TO DEVELOP ADVERSE SIDE EFFECTS
- 11. SUPPRESSION SIDE EFFECTSANTIHISTAMINES ONLY ABOUT 1/3 OF PET PARENTS REPORT SUCCESS TRY SEVERAL DIFFERENT TYPES BEFORE GIVING UP COMMON SIDE EFFECTS CAN RANGE FROM DROWSINESS TO SEIZURES TO NAUSEA.
- 12. SUPPRESSION SIDE EFFECTSCYCLOSPORINE FEW KNOWN SIDE EFFECTS TO DATE, LONG TERM RISKS STILL UNCERTAIN COMMON SIDE EFFECTS INCLUDE: VOMITING, NAUSEA, SOFT STOOLS LESS COMMON HEPATITIS, BONE MARROW SUPPRESSION, SEIZURES HAVE BEEN REPORTED.
- 13. STEROID’S LEGACY 1.SKIN INCREASED SUSCEPTIBILITY TO INFECTION; THINNING OF THE SKIN. 2. ADRENAL CUSHING’S SYNDROME 3. GASTROINTESTINAL VOMITING, DIARRHEA, ULCERS 4. KIDNEY RENAL COMPROMISE, ELECTROLYTE IMBALANCE, INCREASED THURST/URINATION 5. MUSCLE WASTING AND WEANESS, PENDULOUS ABDOMEN 6. PANCREAS TYPE 2 DIABETES
- 14. PRELIMINARY STEPS MEDICALHISTORY AGE OF SYMPTOMS ONSET ROUTE OF POSSIBLE EXPOSURE RELATIONSHIP FOOD/ENVIRONMENTAL EXPOSURE RESPONSE TO ADMINISTERED TREATMENT (HANDOUT)
- 15. PRELIMINARY STEPS CONTINUED…PHYSICAL EXAMINATION TRACK CLINICAL SIGNS (SEASONAL, PERENNIAL) ADDRESS ANY SECONDARY INFECTIONS DISTINGUISH SIGNS FROM OTHER DISEASE INITIAL LABORATORY TESTS IF NECESSARY TO RULE OUT SYSTEMIC CONDITIONS (HANDOUT)
- 16. ALLERGY TESTING WHENTO CONSIDER AN ALLERGY WORKUP : PET PARENT IS COMMITTED TO TREATMENT PROCESS REALISTIC RESULTS GOOD USE OF FUNDS CONFIRM SUSPECTED DIAGNOSIS NOT FIRST LINE OF DEFENSE RULE OUT OTHER IMMUNOLOGICAL ISSUES FIRST
- 17. TYPES OF TESTINGINTRADERMAL SEROLOGICAL
- 18. INTRADERMAL TESTING WEENOFF SUPPRESSION THERAPY (2-6wks) TWO+ VISITS TO DERMATOLOGIST ANIMAL SEDATED INJECT SMALL AMOUNT OF ALLERGEN MEASURE RESULTS http://www.vlsstore.com/Media/PublicationsArticle/Intradermal.pdf
- 19. SEROLOGY SIMPLE BLOODTEST NO NEED TO WEEN OFF SUPPRESSION THERAPY CAN BE DONE IN 1 VISIT, IN CLINIC COST-EFFECTIVE OPTION RISK REDUCTION TEST FOR FOOD REACTIONS
- 20. TYPES OF SEROLOGYWESTERN BLOT RAST ELISA MODIFIED ELISA
- 21. WESTERN BLOT PROCESSProteins are separated by gel electrophoresis. Proteins are transferred to a sheet of special blotting paper called nitrocellulose. The proteins retain the same pattern of separation they had on the gel. The blot is incubated with a generic protein, to bind to any remaining sticky places on the nitrocellulose. An antibody is then added to the solution which is able to bind to its specific protein. The antibody has an enzyme or dye attached. The location of the antibody is revealed by incubating it with a colorless substrate that the attached enzyme converts to a colored product that can be seen and photographed.
- 22.
- 23. WESTERN BLOT CONTINUED…PROS RELATIVELY CHEAP & FAST METHOD DETERMINE AMOUNT & SIZE OF PROTEINS CONS QUESTIONS RAISED ABOUT RELATIVE ABUNDANCE OF PROTEINS WITHIN A TISSUE ELECTROSTATIC MIGRATION VARIATIONS http://teachline.ls.huji.ac.il/72320/methods-tutorial/western/western-pros.html www.abcam.com/technical (other troubleshooting suggestions)
- 24. RAST (RADIOALLERGOSORBANTASSAY) PROCESS USES A RADIOIMMUNOASSAY TO DETECT SPECIFIC IgE ANTIBODIES. THE SUSPENDED ALLERGEN IS BOUND TO AN INSOLUBLE MATERIAL AND SERUM IS ADDED. IF SERUM CONTAINS ANTIBODIES TO ALLERGEN THOSE ANTIBODIES WILL BIND TO THE ALLERGEN. RADIOLABELED ANTI IgE IS ADDED WHERE IT BINDS AND ALL UNBOUND IgE IS WASHED AWAY. THE AMOUNT OF RADIOACTIVITY IS PROPORTIONAL TO THE SERUM IgE FOR THE ALLERGEN.
- 25.
- 26. RAST CONTINUED… PROSIMPROVED SENSITIVITY WITHOUT LOSING SPECIFICITY REPRODUCIBILITY CONS COST AVAILABILITY OF RADIOIMMUNOASSAY POTENTIAL RISK TO USER DUE TO MATERIALS
- 27. ELISA (ENZYMELINKED IMMUNOASSAY) PROCESS INVOLVES AT LEAST 1 ANTIBODY WITH SPECIFICITY FOR A PARTICULAR ANTIGEN. ANTIGEN IS IMMOBILIZED ON A SOLID SUPPORT (MICROTITER PLATE). DETECTION ANTIBODY IS ADDED, FORMING A COMPLEX WITH THE ANTIGEN. BETWEEN STEPS PLATE IS WASHED TO REMOVE ANY PROTEINS/ANTIBODIES NOT SPECIFICALLY BOUND. ENZYMATIC SUBSTRATE PRODUCES VISIBLE SIGNAL.
- 28.
- 29. ELISA CONTINUED… PROSNO RADIOACTIVITY NEEDED SAFE INEXPENSIVE CONS MORE DIFFICULT TO DEVELOP, CONTROL & STANDARIZE HIGHER PROBABILITY OF FALSE NEGATIVES
- 30. MODIFIED ELISA PROCESS:THE PROBLEM WITH A STANDARD ELISA ASSAYS IS THAT IN THE WASHING PROCEDURES, THE ALLERGEN BOUND TO THE PLATE WASHES OFF AND CONSEQUENTLY WHEN THE PATIENT’S ANTIBODY IS ADDED THERE IS NO ANTIGEN THERE, THUS A FALSE NEGATIVE RESULTS. SPECTRUM’S SPOT TEST ADDRESSES THIS BY USING A PATENTED TECHNOLOGY TO COVALENTLY BIND ALLERGEN TO THE PLATE MUCH LIKE Ag IS BOUND TO THE CELLULOSE DISC IN THE RAST.
- 31.
- 32. MODIFIED ELISA CONTINUED…PROS (SPECTRUM) RELIABILITY OF RAST WITH THE ECONOMY OF SCALE OF THE ELISA COVALENT BONDING CONS STILL CONFUSED WITH STANDARD ELISA’S FALSE NEGATIVE RESULTS RELATIVELY NEW TECHNOLOGY, STILL IN REVIEW
- 33. THINGS TO CONSIDERBOTH RAST AND ELISA ARE ESENTIALLY USING THE SAME PRINCIPAL IN THAT THEY BOTH RELY ON AN ANTIGEN ANTIBODY REACTION OCCURING WITH AN ANTIGEN BOUND TO A MATRIX; LIQUID IN RAST, SOLID IN ELISA. SOMETIMES SOURCE1 FINDS ONE OUTCOME AND SOURCE2 ANOTHER. BOTH HOWEVER CAN BE ACCURATE.
- 34. HYPOSENSITIZATION SUCCESS RATESAS HIGH AS 85-90% REDUCTION IN SYMPTOMS FROM DIETARY CHANGE ALONE =40-50% IMPROVED QUALITY OF LIFE FOR PETS PLANT DERIVED SAFE FOR LONG TERM USE LONG TERM ALLERGY MANAGEMENT TOOL NOT CURE
- 35. HYPOSENSITIZATION BEFORE& AFTER
- 36. HOT TOPICS INALLERGY TEWL STUDIES CORELATION BETWEEN TESTING METHODS RUSH IMMUNOTHERAPY
- 37. Q & ASESSION
- 38. SOURCES: SLIDE 3:VIDEO DR. RUFUS RAJADURAI, M.D.; www.rufusrajadurai.wetpaint.com SLIDE 10: SUPPRESSION SIDE EFFECTS DR. MIKE RICHARDS, DVM http://www.vetinfo.com/ceffect.html DR. BARBARA FORNEY, VMD http://www.wedgewoodpetrx.com/learning-center/professional-monographs/ prednisone-for-veterinary-use.html DR. RON HINES, DVM PhD www.2ndchance.info/cushings.htm SLIDE 18: INTRADERMAL TESTING http://www.vlsstore.com/Media/PublicationsArticle/Intradermal.pdf SLIDES 21-32: SERUM TESTING METHODOLOGIES http://en.wikipedia.org/wiki/ELISA http://en.wikipedia.org/wiki/RAST_test http://en.wikipedia.org/wiki/Western_blot http://teachline.ls.huji.ac.il/72320/methods-tutorial/western/western-pros.html
- 39. SOURCES CONTINUED SLIDE34: HOT TOPICS IN ALLERGY TEWL (TRANSEPIDERMAL WATER LOSS http://publik.tuwien.ac.at/files/pub-et_11355.pdf CORELATION BETWEEN TESTING: INTRADERMAL VS SEROLOGICAL http://www3.interscience.wiley.com/journal/120745931/abstract?CRETRY=1&SRETRY=0 RUSH IMMUNOTHERAPY: A SHORTENED SCHEDULE http://linkinghub.elsevier.com/retrieve/pii/S1081120610626427 ADDITIONAL SOURCES: VETERINARY CLINICAL IMMUNOLOGY RICHARD E.W. HALLIWELL PhD DACVD & NEIL T. GORMAN PhD, DACVIM; 1989 MULLER & KIRK’S SMALL ANIMAL DERMATOLOGY, 6 TH EDITION DANNY W. SCOTT, DVM, WILLIAM H. MILLER, JR. VMD., CRAIG E. GRIFFIN, DVM, 2001 THE 5 MINUTE VETERIARY CONSULT CLINICAL COMPANION (SMALL ANIMAL DERMATOLOGY) KAREN HELTON RHODES, DVM; 2002
- 40. LYNDEE KELVER SPECTRUM LABS [email_address] F: WWW.FACBOOK.COM/VETALLERGY T: WWW.TWITTER.COM/VETALLERGY YT: WWW.YOUTUBE.COM/VETALLERGYSPOT LI: WWW.LINKEDIN.COM/LYNDEEKELVER B: VETALLERGYBLOG.COM THANK YOU!
Editor's Notes
- #2 THANK YOU LEARN MORE ABOUT EMERGING TESTING OPTION NOT NEW, GAINING IN POPULARITY EASE OF USE STEROID INDEPENDENT LESS INVASIVE TODAY GOING THRU THESE OPTIONS WHEN PRACTICING WELL VERSED IN OPTIONS AVAILABLE ABLE TO DETERMINE WHICH YOU FEEL IS IN THE BEST INTEREST OF PATIENTS LET’S GET STARTED…
- #3 THESE ARE THINGS GOING TO COVER TODAY BASICS OF ALLERGY SYMPTOMS SUPPRESSION VS TREATMENT WHEN IN PROCESS TO ORDER TESTING TYPES OF TESTING HOT TOPICS IN ALLERGIES FINALLY Q&A SESSION
- #4 START WITH THE BASICS..WHAT IS AN ALLERGY? ANYONE? GREAT VIDEO CLIP THAT GOES OVER INITIAL AND CONTINUED EXPOSURE
- #5 WHAT ARE SOME OF THE SYMPTOMS ASSOCIATED WITH ALLERGIES? SKIN RESPIRATORY DIGESTIVE CONJUCTIVITIS LETS GET INTO A BIT MORE DETAIL ABOUT EACH OF THESE SYMPTOM GROUPS
- #6 WITH SKIN ISSUES YOU CAN HAVE A WIDE RANGE OF SYMPTOMS THESE ARE JUST A FEW (PHOTO) HERE IS AN EXAMPLE OF WHAT IT COULD LOOK LIKE
- #7 NEXT WE GET INTO RESPIRATORY SYMPTOMS LEFT UNTREATED CAN DEV INTO SECONDARY INFECTION SOMETIMESTHEN BE HEARD THRU STETHOSCOPE
- #8 DIGESTIVE SYMPTOMS INCLUDE BE SURE TO RULE OUT IBD IN YOUR INITIAL WORKUPS ARE SITUATIONS WHERE IBD EXIST ALONG WITH ALLERGIES AS WELL (TWO CAN EXASERBATE EACHOTHER)
- #9 HERE IS A VISUAL OVERVIEW OF COMMON SYMPTOMS OF ALLERGY AND WHERE ON THE BODY THESE SYMPTOMS CAN OCCUR
- #10 NEXT WE ARE GOING TO DISCUSS SUPPRESSION VERSUS HYPOSENSITIZATION SUPPRESSION INCLUDES: STEROIDS -REINFORCE MAST CELL WALL REDUCING LIKELIHOOD OF RUPTURE CYCLOSPORINE -SUPPRESS IMMUNE SYSTEM FROM RESPONDING (EFFECTS ALLERGIES AS WELL AS OTHER DISEASE) ANTIHISTAMINES -BLOCK HISTAMINE RECEPTOR SITES, RESULTING IN LESS HYSTAMINE IN BLOOD STREAM & FEWER OUTWARD SYMPTOMS HYPOSENSITIZATION CONSISTS OF -CONTROLLED AMTS OF OFFENDING ALLERGEN -SUBCUTANEOUSLY -INCREASING DOSES AND FREQUENCY -DERIVED FROM PLANTS—SAFE FOR LONG TERM USAGE
- #11 NEXT LETS DISCUSS SIDE EFFECTS OF SOME OF THE MEDICATIONS USED TO SUPPRESS: STEROIDS IN DOGS: MOST DISCUSSED ISSUE: CUSHINGS DISEASE (85% OF TIME CAUSED BY TUMORS IN PITUITARY OTHER 15% MADE UP OF TUMOR ON ADRENAL GLANDS VET. INDUCED (LONG TERM STEROID USE) RESULT OF EXCESSIVE AMTS OF CORTISOL IN BLOOD RELAX LIGAMENTS IN ABDOMEN (CAUSES ENLARGEMENT OF LIVER) DECREASES GROWTH OF HAIR/THINS SKIN DECREASES MUSCLE MASS (WEAK LIMBS); DECREASES PROD OF CONNECTIVE TISSUE THAT STABILIZES JOINTS) PITUTARY: REGULATES HORMONES AND ADRENAL GLANDS ADRENAL: PRODUCES/REGULATES CORTISOL, EPINEPHRINE (ADRENALINE) AND NOREPINEPHRINE) NOREPINEPHRINE IN LOW LEVELS , BELIEVED EQUATE LOW LEVELS OF AROUSAL & INTEREST; PRESENT IN THOSE IN DEPRESSED STATES (HUMANS) BELIEVED TO MAINTAIN BALANCE BETWEEN AGITATION AND DEPRESSION MORE COMMON ISSUES, GOING TO BE: WEIGHT GAIN HYPERACTIVITY PANTING DIARRHEA DEPRESSION (DUE TO POTENTIAL EFFECT ON ADRENAL FUNCTION) CAN DEVELOP INTO SOMETHING MORE SERIOUS THAT EFFECTS LIVER (DETOXIFY BODY, PROTEIN SYNTHESIS, CHEM. 4 DIGESTION) PANCREAS (PRODUCES INSULIN, GLUCAGON, DIGESTIVE ENZYMES) DUE TO ITS EFFECT ON PACREAS (INSULIN PRODUCTION); INCREASES POSS FOR DEV OF DIABETES DIABETES: STEROIDS OVER STIM PETUITARY GLAND ADRENAL GLAND TO INCREASE PRODUCTION OF CORTISOL INCREASE BLOOD SUGAR CAUSES PANCREASE TO COMPENSATE BY INCREASE INSULIN PRODUCTION IN CATS: REQUIRE HIGHER DOSES NOT EXACTLY SURE WHY, BELIEVED WAY THEIR BODY PROCESS DRUG (STEROIDS TO USABLE CORTISOL) ALSO BECAUSE OF THIS PROCESSING, LESS LIKELY TO DEV ADVERSE SIDE EFFECTS
- #12 ANTIHISTAMINES: IMPORTANT TO NOTE ONLY 1 IN 3 REPORT SUCCESS WITH THIS METHOD OF TREATMENT FRUSTRATING FOR PET PARENTS MAY BE NECESSARY TO TRY SEVERAL TYPES BEFORE GIVING UP COMMON SIDE EFFECTS: VARY GREATLY DROWSINESS SEIZURES NAUSEA
- #13 CYCLOSPORINE (AKA ATOPICA) RELATIVELY NEW TO VET MARKET (NOVARTIS 2002); USED IN HUMANS SINCE 1980S FEW KNOWN SIDE EFFECTS TO DATE LONG TERM RISKS STILL UNCERTAIN MOST COMMON: VOMITING NAUSEA SOFT STOOL LESS COMMON: HEPATITIS (INFLAMATION OF LIVER) BONE MARROW SUPPRESSION (PRODUCE RED & WHITE BLOOD CELLS PLATELETS. RED-OXYGEN/CARBON DIOXIDE, WHITE: FIGHT INFECTION; PLATELETS: CLOT) SEIZURES
- #14 SO WHAT IS STEROIDS LEGACY? THIS IS A VISUAL OVERVIEW OF SOME OF THE AREAS IN THE BODY THAT CAN BE AFFECTED BY LONG TERM STEROID USAGE. PENDULOUS-SAGGING ABDOMEN.
- #15 ALLERGY TESTING SHOULD BE USED FOR VERIFICATION PURPOSES NOT MENT TO DIAGNOSE IF ALLERGIES ARE PRESENT SO LETS TAKE A STEP BACK FOR A MOMENT AND DISCUSS PRELIMINARY STEPS SOULD TAKE TO DETERMINE WHAT SORT OF ISSUES PATIENT HAS MEDICAL HISTORY: AGE OF ONSET ROUTE OF POSSIBLE EXPOSURE INHALANT, ORAL, CONTACT RELATION TO FOOD OR ENVIRONMENTAL EXPOSURE ACCIDENTAL, SEASONAL, PERENNIAL NATURE/COURSE OF REACTION CONCURRENT EVENTS (ILLNESS, EXERCISE, DRUG INGESTION) FEEDING HISTORY (FOOD INGREDIENTS, TREATS, SCRAPS, ETC) RESPONSE TO ADMINSTERED TREATMENT WHAT DID YOU GIVE VS HOW THEY RESPONDED WHAT HAPPENDED WAS AS RESPONSE FAVORABLE LOOK AT IT FROM A CAUSE AND EFFECT STANDPOINT
- #16 NOW THAT WE’VE GONE THROUGH THE MEDICAL HISTORY LETS LOOK AT THE CLINICAL SIGNS WHEN DO WE SEE AN INCREASE IN SYMPTOMS? (SEASONAL/YEAR ROUND) ARE THERE SECONDARY INFECTIONS PRESENT? WHAT ARE THEY? TREATME ACCORDINGLY LOOK AT THE OVERALL SYMPTOMS COULD IN INDICATE OTHER MORE SEROUS ISSUES REVIEW AND ADDRESS IF NECESSARY FINALLY INITIAL LAB TEST MAY BE NEEDED TO RULE OUT SYSTEMIC CONDITIONS (DISEASE THAT AFFECTS VAROUS ORGANS, TISSUES OR THE BODY AS A WHOLE)
- #17 SO AFTER ALL THESE PRELIMINARY STEPS HAVE BEEN TAKEN OTHER DISEASE/INFECTION ADDRESSED OR RULED OUT WHEN SHOULD YOU CONSIDER AN ALLERGY WORKUP? COMMITTED TO TREATMENT: PARENTS NEED TO UNDERSTAND PROCESS, HAVE REALISTIC RESULTS-OUTCOME YES CAN BE AN INCOME STREAM FOR PRACTICE MAKE SURE NOT AT THE COST OF A CLIENT (GOOD USE OF THEIR FUNDS). ALLERGY TESTING FOR VERIFICATION NOT DIAGNOSTICS, WHAT YOU ALREADY SUSPECT
- #18 EVEN THOUGH I AM HERE TO SPECIFICALLY ADDRESS SEROLOGY WANT TO TAKE A FEW MOMENTS HERE TO ADDRESS THE PROCESS INVOLVED IN INTRADERMAL TESTING
- #19 SO INTRADERMAL TESTING, WHAT ARE THE STEPS? ACCORDING TO THE MATTHEW J RYAN VETERINARY HOSPITAL AT UNIVERSITY OF PENN. WEEN OFF SUPPRESSION THERAPY 2-6 WEEKS ANTIHISTAMINES 10-14 DAYS BEFORE INJ STEROIDS 8-12 WEEKS BEFORE ORAL STEROIDS 4 WEEKS BEFORE TOPICAL STEROIDS 3-4 WEEKS BEFORE ESSENTIAL FATTY ACIDS (FISH OIL) 10-14 DAYS BEFORE 2+ VISITS: TYPICALLY FIRST VISIT BRING THEM UP TO SPEED SECOND VISIT CONDUCT ALLERGY TESTING SEDATION: RISKS INCREASE WHEN SEDATION IS PERFORMED (CAN CAUSE NAUSEA) INJECT SMALL AMOUNTS OF ALLERGEN MEASURE RESULTS HERE IS WHAT RESULTS CAN LOOK LIKE
- #20 NOW LETS COMPARE THAT METHODOLOGY WITH SERUM BASED TESTING OR SEROLOGY BLOOD TEST NO NEED TO WEEN OFF SUPP. THERAPY SINGLE VISIT COST EFFECTIVE OPTION ELIMINATES RISKS ASSOCIATED WITH SEDATION, AS PATIENT IS AWAKE DURING PROCEDURE (SIMPLE BOOD DRAW) WITH THIS METHODOLOGY YOU CAN ALSO TEST FOR REACTION TO PROTEINS OR FOODS (WHILE CONTROVERSIAL OUR VETS ARE HAVING GREAT RESULTS) 40-50% OF TIME ABLE TO BRING BELOW THRESHOLD JUST MY DIETARY MOD. THIS IS WHAT THE 96 ALLERGEN MICROTITER PLATE LOOKS LIKE
- #21 THERE ARE 4 DIFFERENT TYPES OF SERUM BASED TESTING WESTERN BLOT RAST ELISA AND MODIFIED ELISA LETS DISCUSS EACH OF THEM BRIEFLY
- #22 WESTERN BLOT CONSISTS OF THE FOLLOWING PROCESS: PROTEINS SEP BY GEL ELECTOFORESIS THEN TRANSFERRED ON TO A SHEET OF NITROCELLULOSE (PROTEINS RETAIN SAME PATTERN OF SEPERATION HAD ON CELL) THE BLOT IS INCUBATED WITH A GENERIC PROTEIN TO BIND TO ANY REMAINING STICK PLACES ON NITROCELLULOSE AN ANTIBODY IS ADDED, ABLE TO BIND TO ITS SPECIFIC PROTEIN (HAS ENZYME OR DYE ATTACHED) LOCATION OF ANTIBODY IS REVEALED BY INCUBATEING WITH COLORLESS SUBSTRATE (ATTACHED ENZYME CONVERTS TO COLORED PRODUCT) MEASURED (PHOTOGRAPHED)
- #23 HERE IS AN EXAMPLE OF THE PROCESS
- #24 NOW THAT WE KNOW PROCESS WHAT ARE THE PROS AND CONS PROS: RELATIVELY INEXPENSIVE & FAST METHOD DETERMINES AMT & SIZE OF PROTEINS CONS : QUESTIONS RAISED ABOUT RELATIVE ABUNDANCE OF PROTEINS WITHIN A TISSUE ELECTROSTATIC MIGRATION VARIATIONS: TOO FAST, TOO HOT SWAY RESULTS
- #25 NEXT WE MOVE ON TO THE RAST TESTING RAST STANDS FOR RADIO-ALLER-GO-SORBANT ASSAY PROCESS CONSISTS OF: USING RADIO-IMMUNO ASSAY TO DETECT SPECIFIC IgE ANTIBODIES SUSPENDED ALLERGEN BOUND TO INSOLUBLE MATERIAL (SERUM ADDE) IF SERUM CONTAINS ANTIBODIES TO ALLERGEN (ANTIBODIES WILL BIND TO ALLERGEN) RADIO-LABELD ANTI IgE ADDED; WHERE BINDS ALL UNBOUND IgE WASHED AWAY AMT OF RADIOACTIVITY PROPOTIONAL TO SERUM IgE FOR ALLERGEN.
- #26 THIS IS AN ILLUSTRATION OF THE PROCESS WE’VE JUST COVERED
- #27 PROS AND CONS ARE AS FOLLOWS PROS : IMPROVED SENSITIVITY WITHOUT LOSING SPECIFICITY (SHOW MORE CLEARLY REACTION WITHOUT LOSING SPECIFICLY WHAT ITS REACTING TO) REPRODUCIBITY CONS: COST AVAILABLE FO RADIO-IMMUNO ASSAY (POST 9/11 EVEN MORE EXPENSIVE, MORE REGULATED, ETC.) POTENTIAL RISK TO USER DUE TO MATERIALS INVOLVED
- #28 NEXT WE COME TO THE ELISA TESTING OR ENZYME LINKED IMMUNO ASSAY THIS PROCESS INVOVES AT LEAST 1 ANTIBODY WITH SPECIFICITY FOR A PARTICULAR ANTIGETN ANTIGEN IMMOBILIZED ON SOLID SUPPORT (MOSTLY MICROTITER PLATES) DETECTION ANTIBODY ADDED (COVALENTLY LINKED TO ENZYME OR CAN ITSELF BE DETECTED BY SECONDARY ANTIBODY—BICONJUGATION) , FORMING COMPLEX WITH ANTIGEN BETWEEN STEPS PLATE IS WASHED, REMOVE UNBOUND PROTEINS/ANTIBODIES NOT BOUND ENZYMATIC SUBSTRATE PRODUCES SIGNAL THAT’S VISIBLE.
- #29 THIS IS AN VISUAL EXAMPLE OF WHAT THIS PROCESS LOOKS LIKE
- #30 PROS AND CONS PROS: NO RADIOACTIVITY NEEDED SAFE INEXPENSIVE CONS: MORE DIFFICULT TO DEVELOP, CONTROL & STANDARIZE HIGHER PROBABILITY OF FALSE NEGATIVES DUE TO WASHING THROUGHOUT PROCESS
- #31 MODIFIED ELISA PROCESS SAME HOWEVER SPECTRUM LABS ADDRESSES THE BINDING ISSUE BY COATING THE WELLS TO ALLOW FOR A COVALENT BOND BETWEEN PLATE AND ALLERGEN MIMICKING HOW Ag BINDS TO CELLULOSE DISC IN RAST TESTING.
- #32 SAMPLE OF WHAT MIRCOTITER PLATE LOOKS LIKE AFTER PROCESS AND MEASUREMENT
- #33 PROS AND CONS PROS: SPECTRUM COMBINES RELIABILITY OF RAST TEST WITH ECONOMY OF SCALE OF ELISA WELLS COATED TO ALLOW FOR COVALENT BONDING CONS: STILL CONFUSED WITH STANDARD ELISA’S FALSE NEGATIVE RESULTS RELATIVELY NEW TECHNOLOGY (AS FAR AS SCI COMMUNITY CONCERNED); STILL IN REVIEW
- #34 NOW THAT WE HAVE GONE OVER THE DIFFERENT TYPES OF SERUM BASED TESTING FOR ALLERGIES THINGS TO CONSIDER BOTH RAST /ELISA USING ESSENTIALLY SAME PRINCIPAL BOTH RELY ON ANTIGEN ANTIBODY REACTION OCCURING W/ ANTIGEN BOUND TO MATRIX BUT LIQUID IN RAST AND SOLID IN ELISA SCIENCE 1 OUTCOME; OTHERS ANOTHER (GIVE EXAMPLE—FOOD ALLERGY TESTING)
- #35 ONCE TESTING IS DONE HOW DO YOU PROCEED WITH TREATMENT OR HYPOSENSITIZATION? SHARE WITH YOU SPECTRUM’S STORY SUCCESS RATE AS HIGH AS 90% IN PART, SHOW ALL REACTIONS RATHER THAN JUST TOP REACTION MODIFY DIET BASED ON OUR RESULTS TREAT FOR ALL REACTIONS (UP TO 21 IN A SINGLE SET) PARTNERING COMPREHENSIVE TEST WITH COMREHENSIVE, COST EFFECTIVE TREATMENT REDUCTION IN SYMPTOMS FROM DIETARY CHANGE ALONE =40-50% IMPROVED QUAL OF LIFE FOR PETS PLANT DERIVED –PLANT EXTRACT AND PHENOL (ALL TREATMENT CONSISTS OF) SAFE FOR LONG TERM USE PLEASE NOTE: NO CURE FOR ALLERGIES, ONLY MANAGEMENT
- #36 HERE ARE SOME BEFORE AND AFTER PHOTOS OF THE PATIENT’S WE’VE TESTED AND TREATED WITH OUR METHODOLOGY
- #37 OTHER STUDIES AND TOPICS YOU MAY WANT TO READ UP ON IF DERM INTEREST YOU TEWL STUDIES: TRANS EPIDERMAL WATER LOSS (2004) WHILE NOT FULLY SUBSTANTIATED YET, IT IS BELIEVED LINK BETWEEN THIS EPIDERMAL BARRIER DEFECT AND THE LIKELIHOOD OF ITS PRESENCE IN ATOPIC CANINE SKIN. (LENGTH/THICKNESS OF STRATUM CORNEUM LIPID DEPOSITS LOWER IN ATOPIC PATIENTS THAN IN NORMAL SUBJECTS—ALSO TOPIC OF OUR NL THIS MONTH AND IS PRESENT ON OUR BLOG AS WELL: VETALLERGYBLOG.COM) CORELATION BETWEEN TESTING METHODS: FOR FIRST TIME EVER DERMS RECONGIZING SERUM TESTING TO HAVE HIGH CORELATION WITH INTRADERMAL TESTING RUSH IMMUNOTHERAPY: RATHER THAN TAKING 9 MONTHS TO INCREASE TOLLERENCE DONE IN CLINICAL SETTING OVER FEW DAYS, OR SOME CASES HOURS) TO GET THEM TO POINT ON MAINTENANCE DOSING SOURCE FOLLOWS PROTOCOLS AND FINDINGS OF THE FOLLOWING STUDY CONDUCTED AT NORTHWESTERN UNIVERSITY & NW MEMORIAL HOSPITAL STUDY CONDUCTED ON DOG WITH SEVERE RAGWEED AND GRASS POLLEN ALLERGIES OTHER SOURCES ON WEB AS FAR AS PROTOCOL, THIS ARTICLE DOESN’T GIVE THAT CURRENTLY HIGH SUCCESS RATES, HIGH RISKS AS WELL; REFER TO COLLEGUES WHO HAVE USED THIS CONCEPT SUCCESSFULLY AND HAVE DATA TO SUPPORT IT
- #38 THANK YOU FOR TAKING THE TIME TO HEAR ABOUT SEROLOGY ITS EVOLUTION IN THE VETERINARY ALLERGY ARENA OPEN THE FLOOR TO QUESTIONS AT THIS TIME….