serology
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| Why do we need serology tests & what do they do? | Serology tests are used for diagnosing illnesses, diseases, pathogens etc. They can detect antigens or antigen-specific antibodies.
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| 3 types of reagents for serology? | polyclonal antibodies
monoclonal antibodies
anti-globulins
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| What are polyclonal antibodies/how are they made? | subject is given antigen, antibodies from many different classes develop as a result and are specific for the multiple epitopes (antibody attachment points)of the antigen. Blood is then collected and serum separated producing the polyclonal antiserum.
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| what is an anti-serum? | blood serum containing antibodies that are against a specific antigen
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| what is an anti-globulin? | immunoglobulin (serum plasma proteins that function as antibodies) from one species is given to another species (becomes that species antigen), that species then makes antibodies in response or antiglobulins which is made into an antiserum by purification
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| What are monoclonal antibodies/or how are they made? | Where a normal cell and myeloma cell combine forming hybridomas, the hybridoma that can produce antibodies specific for the antigen are kept-they go to a serum free medium and produce antibodies that are purified.
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| What is an ELISA? | Enzyme linked immunosorbent assay
used for detecting antigens or antibodies
rapid and sensitive
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| Indirect ELISA? | antigen in wells
blood serum added and antibodies specific bind
serum and antibodies that don't bind are washed away
antiglobulin conjugate with enzyme is added which binds to antibodies
enzyme substrate added that binds to enzyme and produces colour
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| what does an indirect ELISA test for? | antibodies
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| Sandwich ELISA? | tests for antigens
antibody to well, then add antigen, then add 2nd antibody, then add antiglobulin conjugate with enzyme, then add enzyme substrate and test for colour
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| 2 tests of ELISA used in vet clinics? | immunofiltration (snap test)
immunochromatography
both test for antigens
chromate one - 2lines= +ve, 1-line= -ve
filtration- circle in west side-high antigen, circle in east side-low antigen
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| Precipitin reaction tests? | ELISA replaced these, gel diffusion assay and immunoelectrophoresis , for gel positive if bands near-indicates antigen cant get in
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| Agglutination Assays | antibody and antigen react and clump
, bacterial-passive-haemagglutination and complement fixation tests
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| bacterial agglutination | detect antibodies against bacterial antigen, as antibodies will prevent clumping
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| passive agglutination | antibodies/antigen are linked with an inert particle that mixes with antigen/antibody and look for agglutination
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| haemagglutination/ haemagglutination inhibition | viruses agglutinate RBC, but antibodies can inhibit this from happening, very cloudy if agglutination
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| Titration of Antibodies-what do we use it for? | -measures amount of antibodies
-highest dilution before agglutination is a titre
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| Other reasons for testing antibody positive? | past exposure
vaccination
maternal antibodies
cross reactivity to related organisms
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| Other reasons for testing antibody negative? | immunosuppression
blood sample taken too early in disease course
presence of interfering antibodies
congenital persistent infection
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| disease course | takes 2-3wks from signs of disease for an antibody to develop
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| Interpreting test results, how many samples should you take and when? | 2 samples, 1-acute stage and 2-convalescent stage (2-3wks after disease), if 4-fold increase in titre infection did occur
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| Sensitivity? | proportion that test positive
= infected with a +ve test/no of infected
if high means that few false negative results and so nearly all infected are detected as positive
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| specificity? | =proportion non-infected that test negative
=proportion non-infected that test negative/ non-infected
high-few false positive results and all that do not have an infection do test negative
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| Are Serological tests absolutely sensitive and specific? | no, and that is why do multiple different types of tests to deduct conclusions
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