Serological Procedures 6.2
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Serological Procedures 6.2
Serology serological procedures NSHS MLT
serological procedures 6.2
The ability of a particular antibody to combine with one antigen instead of another.
fit between the antigenic determinant and the combining site of the antibody.
some of the determinants of an antigen are shared by apparently unrelated molecules.
strength of a multivalent antibody to bind to a multivalent antigen
the bond between a single antigenic determinant and an individual combining site of an antibody.
excessive amount of antibody that can give a false negative result.
decreased amount of antibodyt caused by an excess of antigen
point of equal antigen and antibody amounts which yield the greatest reaction.
zone of equivalence
deviance from the optimum pH of ___ will alter reactivity of the antibody.
different classes of antibodies react at different __________.
what is the antibody that reacts at warm temperatures?
what is the antibody that reacts at cold temperatures?
incubatin time varies with the _____ of antibody.
which types of antibodies will be all the same class and have only one type of heavy and light chain?
these are uniform, highly specific antibodies that caon be produced in large quantitites, cloned from a single cell to bind to a single specific antigen.
what are the three uses of monoclonal antibodies?
typing erythrocyte antigens
typing leukocyte antigens
as a reagent
serological samples should be centrifuged for __ minutes at moderate speed.
if testing cannot be done promptly the serum should be frozen at what temperature?
how is complement inactivated?
heat serum to 56
C for 30min
if serum is not used within four hours, reheat serum at 56
C for ___ minutes.
the highest dilution in which a reaction occurs.
what is the purpose of a titer?
to determine the concentration of antibody
this is used to make serum less concentrated.
each succeeding dilution is the same as the preceding dilution
calculations of any given dilution can be determined by using this formula:
in the formula D1(V1)=D2(V2) what is the starting dilution.
in the formula D1(V1)=D2(V2) what is the starting volume?
in the formula D1(V1)=D2(V2) what is the final dilution
in the formula D1(V1)=D2(V2) what is the final total volume?
pipettes should be periodically calibrated for what three things?
Red Blood cell Suspensions should be used in serological tests as an ________.
in an RBC suspension what should be used as the diluent?
RBC's contain _______; serve as receptor sites for antibodies or complement.
what are the four carriers used in serology?
what is the calucation that can be used to determine the minimum packed cell volume needed to make a solution?
PCV= (total volume X % desired)/100
in an RBC suspension this represents the packed RBCs.
true or false
Hemolyzed RBC's can be used in an RBC suspension.
Mono and ASO use what carrier?
Floculation tests use what carrier?
what carrier is used in hemaglutination?
in what testing method, if antigens are present in serum, they will bind with antibodies forming visible cross-linked aggregates.
in latex agglutination antibody molecules bind to latex _____.
what are the five latex agglutination immunologic assays?
IgG and IgM rheumatoid factors
in what test are latex particles coated with strptolysin O antigen?
in Antistreptolysin O visible agglutination will be seen mixed with patient's serum containing what?
titers begin to rise at __ days after the onset of infection and peaks at ____ weeks after infection.
In ASO, a four fold increase between acute and convalescent tube indicates a recent infection with what?
Group A strep
ASO antibodies are elevated in acute/convalescent sera with what disease?
acute rheumatic fever
serological testing should compare acute and convalescent sera collected ______ weeks apart.
how long are ASO samples stable at 2-8
how long are ASO specimens stable at -25
this is based on agglutination of horse RBC's by hetrophil antibody present in infectious mononucleosis.
in monotest horse RBC's exhibit antigens against both __________ and ______________ threfore a differential absorption of patient serum is necessary.
the monotest immunologic theory is based on __________ principle.
in the monotest immunologic theory, what contains only forssman antigen which will absorb forssman heterophil antibody?
guinea pig kidney
in monotest immunologic theory what will absorb only no-forssman heterophil antibody of infectious mononucleosis?
in a monotest immunologic theory this is tested with both the guniea pig-serum specimen and the beef RBC-serum specimen.
to be positive for IM heterophil antibody, the guinea pig should be _________ for agglutination and the beef RBC should be _________ for agglutination.
what is the sample for Monotest?
Serum or plasma mixed with EDTA
what is the confirmatory test for infectious mononucleosis?
Epstein-Barr virus antibody
this is the causitive agent of infectious mononucleosis?
this is a DNA herpes-type virus that infects the B-lymphocytes?
this is tested to confirm infectious mononucleosis on immune suppressed patients.
this test is beneficial in defining immune status and stage of infectious mononucleosis.
antigen-antibody __________ when combined in proportions at or near equivalence.
in what type of test is interaction of soluble antigen with antibody that results in the formation of a precipitate of fine particles?
in flocculation test particles are ____________ or _____________ visible.
what are two examples of flocculation tests?
this test is based on agglutination of RBC's with antigens that are natural or bound to RBC's
what are three examples of hemagglutination immunologic assays?
Precipitation methods are performed in what two mediums?
semi solid media (agar or agarose)
non-gel support medium (cellulose acetate)
what are the two types of precipitation methods?
in this test antibody dilutions and specific antigens are placed in adjacent wells. Antigen and antibody diffuse out, bind together to form a visible precipitate.
what are the three basic reaction patterns in double immunodiffusion?
In which double immunodiffusion result in precipitin band forms a single smooth arch?
in which double immunodiffusion resuld do precipitin lines merge in a spur formation?
in which double immunodiffusion reslut do precipitin lines cross each other?
what are the acceptable specimens for double immunodiffusion?
in double immunodiffusion why are urine specimens tested in concentrated and unconcentrated forms?
wide range of light cahin concentrations
what are the two double immunodiffusion immunologic assays?
systemic lupus erythrematosus
Radial immunodiffusion (RID) is used to identify and quantitate the number of _________ found in human serum.
in radial immuno diffusion (RID) the antigen will diffuse throughout the agan and form what?
a precipitin ring
in radial immunodiffusion specific antibody is added to a buffered agarose medium, test ________ is added to a well in the agarose.
in radial immunodiffusion (RID) diameter of the precipitin rig is related to what?
what are the four clinical applications of radial immunodiffusion (RID)?
what are the two electro immunodiffusion methods?
Countercurrent immunoelectrophoresis (CIE)
in what test do antigens and antibodies move more quickly toward each other with addition of electrical current?
countercurrent immunoelectrophoresis (CIE)
in CIE agarose gel is used. pH is such that antibodies are ________ charged and antigens are ________ charged.
in countercurrent immiunoelectrophoresis how is the zone of equivalence represented?
antigen-antibody complex forsm a visible line of precipitin
what is the clinical application of countercurrent immunoelectrophoresis (CIE)?
detection of anti-nuclear ribonucleoprotein (nRNP):
associated to sharp's syndrome (mixed connective tissue disease)
what test is associated with sharp's syndrome (mixed conective tissue disease)?
detection of anti-nuclear ribonucleoprotein (nRNP)
in rocket electrophoresis is based on antigen ______________ in an antibody containing gel.
in rocket electrophoresis what is adjusted to inhibit antibody migration?
in rocket electrophoresis the height of the rocket is __________ to the antigen concentration.
what ar the four types of labeling techniques?
radio immuno assay (RIA)
enzyme immunoassay (EIA)
what are the two assays that involve chemiluminescence?
what are the two types of immunofluorescent techniques?
in what test are radioisotopes used to measure the concentration of antigens or antibodies
in what test are radiolabled ligands bound to antigen-antibody complex?
in RIA radiactivity is counted on what?
this test is extremly sensitive and able to detect trace amounts of antigen or antibody.
what are the two clinical applications of radioimmunoassay (RIA)
hepatitis A IgM antibody
hepatitis A antigen
in this test lables can be attached to antigen or antibody.
in this chemiluminescence assay, a fixed amount of labeled antigen competes with unlabeled antigen (patient sample) for a limited numberr of antibody-binding sites.
in a competitive assay the amount of light emitted is _________ proportional to the amount of analyte (antigen) being tested.
in this chemiluminescene test sample antigen binds to a fixed antibody on a solid phase, then a second antibody labled with chemiluminescent binds to the antigen-Ab complex on the solid phase.
in a sadwich assay the amount of light emitted is __________ proportional to the amount of the analyte (antigen) being tested.
what are the two clinical applications of chemiluminescence?
hepatitis A IgM antibody
hepatitis A antigen
this test is similar to RIA but used a non-isotopic label.
this test is safer than RIA yet has the same specificity and sensitivity as RIA.
enzyme immunoassay (EIA)
in Enzyme immunoassay (EIA) what is used instead of a radioisotope?
an enzyme labeled antibody or antigen conjugate
what ar the most commonly used enzymes to detect the presence and quantity of antigen or antibody in patient serum?
in EIA substrate causes color production and is measured how?
what are the three clinical applications of Enzyme Immunoassay (EIA)?
in this extremely specific and sensitive test, fluorescent molecules are used instead of radioisotopes or enzyme labels.
what are the three clinical applications of immunofluorescent techniques?
in this immunofluorescene technique conjugated reagent antibody is used to detect antigen-antibody complex at microscope level.
in direct immunofluorescence what is used ot visualized many bacteria in direct smears?
fuorescein-conjugated antibodies boud to fuorochrome
direct immunofluorescence uses the immunological/physiological theory of:
intracellular antigen detection
what are the two sepcimens used for direct immunofluorescence?
in this test, antigen source such as toxoplasma microorganism or virus of infected tissue culture cells to the specific antibody being tested is affixed to a slide.
in indirect immunofluorescence what reacts with antigen forming an antigen-antibody complex?
unkown antibody (patient's Ab)
in indirect immunofluorescence what is added to the Ab-Ag complex to bind to the antibody as a marker?
labeled anti-immunoglobulin (fluorescent Ab)
indirect immunofluorescence measures what?
direct immunofluorescene measures what?
what is the specimen collected for indirect immunofluorescence?
what are three indications for performing indirect immunofluorescene on serum?
virus infected tissue culture cells
what is the classic method for demonstrating presence of a complement fixing antibody in serum?
what is the indicator system for complement fixation?
complement from guinea pig serum
in complement fixation patient serum is added to _______ antigen, and complement is added to the solution.
in complement fixation, if serum contains the antibody for the antigen, then an antigen-antibody complex will bind with what?
in complement fixation sheep RBC's with anti-sheep antibody are added to the solution of antibodies, antigen, and complement. if complement has not been bound, it is available to bind with what?
coated sheep RBC's
hemolysis of indicator sheep cells indicates a lack of antibody in patient serum and a _________ complement fixation test.
how is a positive complement fixation test demonstrated.
lack of hemolysis
why is a positive complement fixation test demonstrated by the lack of hemolysis in indicator sheeps RBC's?
patients serum does contain a complement fixing antibody
what are the three clinical applications of complement fixation?
Herpes simplex infection
this is an in vitro method that amplifies low level of specific DNA sequences in a sample to higher quantities.
polymerase Chain Reaction
in PCR ___ short DNA primers are used as templates that follow the enzymatic process.
in PCR the enzymatic prcess is carried out in cycles and repeated up to ___ times.
what are the four steps of enzymatic process in PCR?
Extension of primed DNA sequenced
The amount specific DNA sequence double and examined in various ways
polymerase chain reaction is used in the detection of ____.
what are the two clinical applications for PCR?
human papilloman virus (HPV)
coronary artery disease
If the nucleic acid of interest is RNA rather than DNA, the PCR procedures can be modified to include the conversion of RNA to DNA using what?
Reverse Transcriptase-Polymerase (RT-PCR)
the clinical application for RT-PCR is to test for what?
what are the two clinical applications of RNA?
Human Immunodeficiency Virus
Hepatitis C Virus