to detect antigens and antibodies that may be mall in size or present in very low concentrations using a labeled reactant to monitor the specific antigen-antibody binding
what is the process for competitive immunoassays
all reactants are mixed together simultaneously
labeled antigen competes with unlabeled patient antigen for a limited number of antibody binding sites
the amount of bound label is inversely proportional to the concentration of the labeled antigen
competitive immunoassays
describe process of noncompetitive immunoassays
antibody is anchored and absorbed into a solid phase
unknown patient antigen is reacted an captured by antibody
Solid is washed to remove unbound antigen and an anti antibody with label is added to the reaction
noncompetitive immunoassays concentration of labelled and unlabelled antibody
the amount of label measured is directy proportional to the amount of patient antigen
non competitive immunoassay
theory behind Enzyme immunoassays (EIAs or ELISA)
using enzymes as labels which react with substrates to produce breakdown product that may be fluorogenic, luminescent, or chromogenic
what is the most commonly used enzyme label
alkaline phosphatase
horseradish peroxidase
both have high sensitivity and easy to detect
highest conversion of substrate rates (turnover)
what are ELISAs used for
HIV
hepatitis B
ELISAs produce a sandwhich immunoassays because
antigens that have multiple determinants such as
antibodies
cytokines
tumor markers
heterogeneous ELISA or homogeneous ELISA require washing step
heterogeneous
which is more sensitive
heterogenous assay or homogenous assay
heterogenous assay
homogenous EIAs
do not require a washing step
enzyme activity is proportional to concentration of patient antigen present
fluorescent immunoassays are restricted in what way
qualitative observations using a fluorescent microscope
fluorescent immunoassays are used for
identification of microorgansism in cell culture or infected tissue, neoplastic tissue
used to determine concentration of therapeutic drugs
described the theory of chemiluminescent immunoassays
involves the emission of light caused by oxidation reaction producing an excited molecule as it decays back to its original ground state
can be used for heterogeneous and homogeneous assays because labels can be attached to either antigen/ antibody.