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56 Cards in this Set
- Front
- Back
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the initial force of attraction of an antibody for an antigen
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affinity
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strength of binding; the force that keeps the molecule together once it has formed
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avidity
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they key portion of the immunogen against which the immune response is directed; also known as the determinant site
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epitope
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when antibodies attach to antigens that are similar to but not the same as the antigen for which it was originally produced
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cross reactivity
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the process of measuring the loss of intensity of transmitted light due to the scattering effect of particles suspended in it; antigen is mixed with its specific antibody, the immune complex is formed, lattice formation takes place, the molecule becomes big enough to fall out of solution, and the solution which was once clear, will now become turbid, or cloudy; measures unscattered light
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turbidimetry
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How do we use turbidimetry to find the unknown?
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the cloudiness of the solution can tell us how much of the unknown (be it the antigen or the antibody) is present; we can use SPECTROPHOTOMETRY to measure the amount of turbidity present by measuring the decreased amount of light passing through the solution; we are looking for an immune complex formation
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formed from the integral binding of an antibody to a soluble antigen
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immune complex
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when there is TOO MUCH ANTIBODY, there are not enough to form cross-links between the antigens and so no lattice formation; this means that the patient has huge amounts of antibody, but we can't detect them because there are too many of them; this RESULTS IN A FALSE NEGATIVE THAT IS CAUSED BY TOO MUCH ANTIBODY
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prozone phenomenon (or prozone reaction)
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when there is TOO MUCH ANTIGEN, each antigen is bound to each antibody binding site; cross links cannot be formed between the antibodies, so no lattice formation and no precipitation; this RESULTS IN A FALSE NEGATIVE CAUSED BY TOO MUCH ANTIGEN
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postzone phenomenon (or postzone reaction)
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involves combining soluble antigen with soluble antibody; when the two dissolved substances combine to form an immune complex, the immune complex formed is so big that the molecule falls out of solution; the solution changes from clear to cloudy
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precipitation assay
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principles use particles that will clump together when an immune complex is formed; particles used can be bacteria, red cells, or latex particles, to name a few
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agglutination
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give examples of a precipitation test
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(?) TURBIDIMETRY, nephelometry
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give examples of an agglutination test
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(?) Widal Test (looks for the detection of antibodies occurring in typhoid fever, brucellosis, and tularemia), direct, passive, reverse passive, agglutination inhibition, coagglutination
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What kind of agars do we use when we use electrophoresis?
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support medium is a gel (either ARAGOSE or AGAR)
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What is the difference between aragose and agar?
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agar has a strong negative charge; agarose has almost none
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Which electrophoresis support medium is preferred over the other and why?
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agarose is preferred to agar because agar has a strong negative charge and agarose has almost none
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a more sensitive way to measure precipitation; some particles will cause light to be deviated (reflected) at very specific angles; measures the ability of a solution of immune complexes to scatter the light and uses that to measure the number of complexes present, and so the amount of the unknown that is present; measures scattered light
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nephelometry
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What is an example of something that we test often using nephelometry?
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(?) serum proteins, quantification of immunoglobulins such as IgG, IgA, IgM, and IgE and kappa and lambda light chains; complement components, C-reactie protein, several clotting factors
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What type of test is rocket electrophoresis?
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precipitation
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single diffusion method; the precipitin line that is formed is conical in shape, resembling a rocket
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Rocket Immunoelectrophoresis
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double diffusion method; both antigen and antibody diffuse independently through a semi-solid medium in two dimensions, horizontally and vertically; reactants are added to wells cut in the agar; white lines of precipitation are formed where equal amounts of antigen meet equal amount of antibody; arc formation indicates that the two antigens are identical; two crossed line indicates the antigens share no identical determinants; partial identity indicates that one antigen shares a determinant with the other antigen, the spur always points to the simpler antigen
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Ouchterlony Double Diffusion
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Give an example of a single diffusion method.
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RADIAL IMMUNODIFFUSION (might also be rocket immunoelectrophoresis)
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What is the zone of equivalence?
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where the antigen and the antibody are at equal amounts, which causes an immune complex to form and precipitate
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What do we call the antibodies that cause agglutination?
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agglutinens
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What is batching?
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(?)
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What is the optimal pH for most antigen antibody reactions?
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(?) 8.6
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IgG reacts best at what temperature?
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(?) warm
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IgM reacts best at what temperature?
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(?) cold
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systems using bacteria as the inert particles to which antibody is attached (S. aureus is most frequently used because it has a protein on its outer surface called protein A, which naturally adsorbs the Fc portion of antibody molecules
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coagglutination
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occurs when antigens are found naturally on a particle
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direct agglutination
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uses haptens that are complexed to proteins; NO AGGLUTINATION IS A POSITIVE REACTION
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Agglutination Inhibition
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uses particles that have the antigen put on them; the antigen is not there naturally
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passive or indirect agglutination
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antibody is attached to the carrier particle
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reverse passive agglutination
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red cells are the indicator particles
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Hemagglutination Inhibition
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Give an example of a "complete" antibody.
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IgM
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Give an example of an "incomplete" antibody.
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IgG
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Why do we call antibodies complete or incomplete?
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complete antibodies can agglutinate on their own without enhancements (such as warming or centrifuging); incomplete antibodies need enhancement techniques to cause agglutination
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an antibody specific for the analyte we are searching for is bound to a solid phase vehicle, which may be a test tube or a microtiter well; patient sample is mixed with the antigen (analyte) which has been tagged with a radioisotope; this mixture is then added to the solid phase; if patient antigen is present, some of the binding sites will be filled with unlabeled analyte, thus decreasing the amount of bound radioactive labeled antigen; excess is rinsed or washed away, and the amount of radioactivity in the test tube is measured; the amount of label in the bound phase is indirectly proportional to the amount of patient antigen present
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competitive binding
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What is another name for sandwich immunoassay?
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capture assay (we did one in the lab for strep)
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When was the immunoflourescent assay technique first used and for what?
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(?) it was first used to find certain antigens located in tissues; is is used for syphilis
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What is ELISA and how does it work?
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Enzyme-Linked Immunosorbent Assays; the old method used competitive binding where enzyme-labeled antigen competes with unlabeled patient antigen for a limited number of binding sites on antibody molecules that are attached to a solid phase; enzyme activity is measured by adding a chromogen (substance that will cause a color reaction wherever there is enzyme activity); most labs now use non competitive assays with labeled enzymes because they are eve more sensitive
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If we get a positive RPR for syphilis, we must send it out for a _ test. A positive on that test is _.
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IFA (immunoflourescent assay) , diagnostic
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What is the special light for IFA (Immunoflourescent Assay)?
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tungsten halogen or mercury vapor arc lamp
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the emission of light caused by a chemical reaction , typically an oxidation reaction, producing an excited molecule that decays back to its original ground state
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Chemiluminescence
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when DNA strands spontaneously join back to the strand that it matched originally after heat is removed
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anneal
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binding of two complementary strands of DNA
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hybridization
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when the double helix of the DNA molecule is easily separated by exposing the DNA to high temperatures or an alkaline solution
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denaturation
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a short strand of DNA or RNA of a known sequence that is used to identify the presence of a complementary single strand of DNA in a patient specimen
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nucleic acid probe
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How many nucleic acid bases do there need to be in consecutive order to detect the hybrid?
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16-20
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clinical samples are applied directly to a membrane surface; membrane is heated to denature the DNA strands and then labeled probes are added; the membrane is ashed to remove any unhybridzed probe and the presence of remaining probe is detected by a method that will detect the label used
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dot-blot assay
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uses two probes and allows the sample to be more purified; a capture probe is attached to the membrane and then the labeled probe is added after the patient sample has been allowed to anneal
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sandwich hybridization
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analysis of DNA fragments using probes in a solid phase
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Southern blot
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used to extract a separate RNA
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Northern blot
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can take place when both the target nucleic acid and the probe are free to interact in a reaction mixture
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solution hybridization
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a type of hybridization reaction in which the target nucleic acid is found in intact cells
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Situ Hybridization
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know western blot
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uses immunofixation electrophoresis
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