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IMMUNO AND SERO
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MIDTERMS
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Labeling techniques in immunoassay
level: Labeling techniques in immunoassay
Questions and Answers List
level questions: Labeling techniques in immunoassay
Question
Answer
2 TYPES OF IMMUNOASSAY FORMAT
1. HETEROGENOUS 2. HOMOGENOUS
INVOLVES A SOLID PHASE (MICROWELL AND BEADS) AND REQUIRE WASHING STEPS TO REMOVE UNBOUND ANTIGENS OR ANTIBODY
HETEROGENOUS
ONLY CONSIST OF LIQUID PHASE AND DO NOT REQUIRE WASHING STEPS. RELATIVELY EASIER AND FASTER.
HOMOGENOUS
TYPES OF IMMUNOASSAY
ENZYME IMMUNOASSAY CHEMILUMINESCENCE ELECTROCHEMILUMINESCENCE FLUOROIMMUNOASSAY
DESIGNED TO DETECT ANTIGEN OR ANTIBODIES BY PRODUCING AN ENZYME-TRIGGERED COLOR CHANGE
ELISA- ENZYME-LINKED IMMUNOSORBENT ASSAY
USES A NONISOTOPIC LABEL THAT OFFERS SAFETY WHICH IN THIS CASE, AN ENZYME:
HIGH DEGREE OF STABILITY EXTREME SPECIFICITY ABSENCE FROM AG OR AB NO ALTERATION BY INHIBITOR WITHIN THE SYSTEM
AN ANTIGEN SPECIFIC ENZYME IS ATTAHCED TO A _____SURFACE
SOLID PHASE
SAMPLE WHICH MAY CONTAIN THE ANTIGEN IS ADDED ENZYME LABELLED ANTIBODY (CONJUGATE) IS ADDED A CHROMOGENIC SUBSTANCE IS ADDED TO HAVE FINAL RESULT WHICH IS COLOR CHANGE
ANTIGEN DETECTION
ANTIBODY IS DETECTED DIRECTLY PROPORTIONAL TO A COLOR CHANGE
NONCOMPETITIVE ENZYME ASSAY
ANTIBODY IS DETECTED INVERSELY PROPORTIONAL TO THE COLOR CHANGE
COMPETITIVE ENZYME ASSAY
SPECIFIC ANTIBODY AGAINST SPECIFIC DISEASE IS BEING MEASURED
CAPTURE ENZYME IMMUNOASSAY
REFERS TO THE LIGHT EMISSIION PRODUCED DURING A CHEMICAL REACTION
CHEMILUMINESCENCE
MOST CHEMICALS USED IN THIS METHOD ARE STABLE AND NONTOXIC
CHEMILUMINESCENCE
COMPETITION BETWEEN LABELLED ANTIGEN AND PATIENT ANTIGEN (INVERSE)
COMPETITIVE IMMUNOASSAY
LABELLED ANTIBODY ATTACH TO AG-AB COMPLEX WHICH LIGHTS UP.
SANDWICH IMMUNOASSAY
DETECTS LIGHT EMITTED
PHOTOMULTIPLIER TUBE
ENZYME OR RADIOISOTOPE ARE REPLACED BY FLUORESCENT MOLECULE
IMMUNOFLUORESCENCE
FLUORESCENT COMPOUND WITH HIGH AFFINITY TO PROTEINS AND AG-AB COMPLEXES
FITC(FLUORESCEIN ISOTHIOCYANATE-
DIRECT ADDITION OF FLUORESCEIN
DIRECT IMMUNOFLUORESCENT ASSAY-
INCLUDES WASHING
INHIBITION IMMUNOFLUORESCENT ASSAY
FOLLOWS THE THEORY THAT ANTIBODY CAN ALSO BE AN ANTIGEN. WIDELY USED IN DETECTION OF DIVERSE ANTIBODIES
INDIRECT IMMUNOFLUORESCENT ASSAY-
USE OF SEMICONDUCTOR NANOCRYSTAL AS A FLUORESCENT LABELING ANTIBODIES
QUANTUM DOTS (Q DOTS)
SUPERCONDUCTING QUANTUM INTERFERENCE DEVICE (SQUID) DETECTS ANTIBODY THAT IS TAGGED TO ANTIGEN THROUGH SUPERMAGNETIC PARTICLE
SQUID TECHNOLOGY
MOVEMENT OF SINGLET OXYGEN THROUGH THE USE OF LIGHT ENERGY
LUMINISCENT OXYGEN-CHANNELING IMMUNOASSAY (LOCI)
TYRAMIDE SIGNAL AMPLIFICATION(TSA) DETECTS B-CELL CLONALITY IN TISSUE SPECIMENS (IMMUNOHISTOPATH)
SIGNAL AMPLIFICATION TECHNOLOGY-
USED IN AUTOMATED DNA SEQUENCES IN DNA DETECTION, SHORTER TURN AROUND TIME
MAGNETIC LABELING TECHNOLOGY-
A TYPE OF ELISA WHICH IS TIME SPECIFIC TO EXCLUDE BACKGROUND INTERFERENCE OF LIGHT
TIME-RESOLVED FLUOROIMMUNOASSAY
BASED ON THE RATE OF ROTATION OF CONJUGATE. REDUCED ROTATION MEANS BONDING
FLUORESCENCE POLARIZATION IMMUNOASSAY
USES FLUORESCENT MOLECULES TO BRIGHTLY “PAINT” GENES OR CHROMOSOMES
FLUORESCENCE IN SITU HYBRIDIZATION (FISH)
CHARACTERISTICS OF AUTOMATED TESTING
PREANALYTICAL ANALYTICAL POSTANALYTICAL HARMONIZATION
LABELLING, ACCESSIONING AND TRACKING
PREANALYTICAL
AUTOMATED RESULTS ENTRY, QC, VALIDATION, LIS
ANALYTICAL
PATIENT REPORTS, RECORDS, NETWORK TO OTHER SYSTEM
POSTANALYTICAL
PROCESS IN TAKING STEPS TO ACHIEVE UNIFORMITY OF RESULTS BY DIFFERENT GROUPS
HARMONIZATION
COMPLEXES FORMED FROM SERUM (ANALYTE) AND REAGENT WILL ALLOW THE LIGHT TO SCATTER WHEN PASSED THROUGH THE SOLUTION. THEREFORE, NEPHELOMETRY IS THE MEASUREMENT OF LIGHT SCATTERED
NEPHELOMETRY
DETECTS THE LIGHT SCATTERED PHOTOMETRICALLY
PHOTODIODE
PEG
POLYETHYLENE GLYCOL- ACTS AS STABILIZER OF THE SOLUTION
THE AMOUNT OF SCATTERED LIGHT IS _______ TO THE AMOUNT OF INSOLUBLE COMPLEXES.
DIRECTLY PROPORTIONAL
A RELATIONSHIP BETWEEN THE QUANTITY OF ANTIGEN AND MEASURING SIGNAL AT A CONSTANT ANTIBODY CONCENTRATION.
HEIDELBERGER CURVE
LIGH IS SCATTERED IN VARIETY OF ANGLES- SIDE, FORWARD AND BACK SCATTER
PHYSICAL BASIS
HIGH PERFORMANCE LIGHT EMITTING DIODE IS USED AS A LIGHT SOURCE (840nm) AND SILICON PHOTODIODE AS DETECTOR
OPTICAL SYSTEM
EMPTY CUVETTE IS USED AS BLANK, CALCULATION IS AUTOMATICALLY PERFORMED BY SYSTEM COMPUTER
MEASURING METHOD
RAPID, REPRODUCIBLE AND SIMPLE TO OPERATE
ADVANTAGE
HIGH MATERIAL COST, INTERFERRING SUBSTANCE SUCH AS BACTERIA THAT MAY CAUSE PROTEIN DENATURATION AND LIPEMIA THAT MAY EXCEED THE PRESET LIMITS.
DISADVANTAGE
THESE ARE PROTEINS THAT PRECIPITATE OR GEL AT COLD TEMPERATURE. CAUSES COLD PRECIPITATES DURING BLOOD TESTING.
CRYOGLUBULINS
IN 1917, EINSTEIN SPECULATED THAT UNDER CERTAIN CONDITION, LIGHT COULD BE ABSORBED BY ATOMS THEN BE STIMULATED TO SHED ITS GAINED LIGHT. LASER HAS NOW BEEN WIDELY USED BOTH INDUSTRIAL AND MEDICAL APPLICATIONS.
FLOW CELL CYTOMETRY
LIGHT AMPLIFICATION BY STIMULATED EMISSION OF RADIATION. A CONCENTRATED BEAM OF LIGHT COMPOSED OF ONE OR UNIFORM WAVELENGTH
LASER
BASIC UNIT OF ALL RADIATION
PHOTON
BASED ON STAINING CELLS WITH FLUOROCHROME DYES THAT EXCITES AND FLUORESCE WHEN PASSED WITH LASER LIGHT THAT IS DETECTED BY A PHOTODETECTOR. THEREFORE, FLOW CELL CYTOMETRY IS THE MEASUREMENT OF LIGHT PRODUCED BY THE REACTED CELLULAR COMPONENT
FLOW CELL CYTOMETRY
ACRIDINE ORANGE AND THIOFLAVIN T.
DYES USED
GAS LASER, KRYPTON
LASER USED
IMMUNOPHENOTYPING IS THE MOST COMMONLY USED METHOD FOR FLOW CELL CYTOMETRY. DETECTS DIFFERENT TYPES OF CELL BY THEIR CD
CLINICAL USE
USES 4 LASER LIGHT OFTEN PRODUCES 16 DIFFERENT COLORS. OFFERS GREAT SENSITIVITY AND SPECIFICITY
MULTICOLOR IMMUNOFLOURESCENCE
WHOLE BLOOD, ASPIRATES, BONE MARROW CAN ALL BE USED IN FLOW CELL CYTOMETRY. EDTA IS THE ANTICOAGULANT OF CHOICE.
SPECIMEN