Inert particles are sensitized with either antigen or antibody. Which of the following tests is used extensively to detect microbial antigens rapidly (five minute or less)?
|C||Enzyme-linked immunosorbent assay (ELISA)|
a. Of the many methods available for antigen and antibody detection, LA, ELISA, RIA, CIE, and COA are the most widely used.
b. Latex agglutination (LA) employs latex polystyrene particles sensitized by either antibody or anti gen. LA is more sensitive than CIE and COA but slightly less sensitive than either RIA or EIA. Passive agglutination increases the sensitivity of detection of RA, CRP, ASLO.
c. LA has been used to detect H. influenzae, Neisseria meningitidis, and S. pneumoniae antigens in cerebrospinal fluid.
d. LA has also been used for detection of cryptococcal antigen. Most recently, LA has been widely used for rapid detection of group. A streptococcal antigen directly from the pharynx. The test is rapid (five minutes), sensitive (approximately 90%), and specific (99%).
e. Coagglutination (COA), also an agglutination test, is slightly less sensitive than LA but is less susceptible to changes in environment (e.g., temperature).
f. Most strains of coagulase-positive staphylococci have protein A in their cell wall. Protein A binds the Fc fragment of microbial antigens in body fluids. COA has also been used to rapidly type or group bacterial isolates.
g. Enzyme immunoassays (ETAs) can be either homogeneous (EMIT) or heterogeneous (ELISA). EMIT has been used primarily for assays of low- molecular-weight drugs. Its primary use in microbiology has been for assays of aminoglycoside antibiotics.
h. EIAs vary as to the solid support used. A variety of supports can be used, such as polystyrene microdilution plates, paddles, plastic beads, and tubes.
i. The number of layers in the antibody- antigen sandwich varies; usually as additional layers are added, detection sensitivity is increased. The two most common enzymes are horseradish peroxidase (HRP) and alkaline phosphatase (AP). -galactosidase has also been employed.
j. Orthophenylene diamine is the most common substrate for HRP and p-nitrophenyl phosphate for AP Because EIAs are usually read in the visible color range, the tests can be read qualitatively by eye or quantitatively by machine.
k. Counterimmunoelectrophoresis (CIE) was originally used for “Australia antigen” (HBsAg) but was soon replaced by RIA. For a decade, CIE was used to detect antigens in body fluids. CIE is not an easy technique.
l. Its success depends on the control of many variables, including solid support, voltage, current, buffer, affinity and avidity of antibodies, charge on the antigen, and time of electrophoresing.
m. Radioimmunoassay involves the radiolabeling of either antibody or antigen (Ag) using ‘ (131I) or 125I (radioisotopes). It measures very small quantities and can be used to detect hormones, carcinoembryonic antigen (CEA), hepatitis B Ag, steroids, prostaglandins, and morphine- related drugs in patient sera.