Immunoassays types

These matrices have been used as chromatographic stationary phases for both enantiomeric and stereoisomeric separations [8], as artificial antibodies in immunoassay-type analyses [7], as microreactors for stereoselective synthesis [9] and as catalysts [10,11] and equilibrium-shifters [12,13] for a variety of reactions. Since MIPs can be produced for such a wide variety of molecules, these characteristics have led many researchers to investigate the possibilities of employing MIPs as the... 

Heterogeneous immunoassay Type of immunoassay that requires separation of bound and free fractions... 

Immunoassay Type Label type Label Property Measured... 

The Centers for Disease Control and Prevention (CDC) maintains a Web site to provide information related to AIDS and HIV. Use your Web browser to connect to http //cheiTiistry.brookscole.com/skoogfac/. From the Chapter Resource Menu, choose Web Works. Locate the Chapter 11 section, and click on the link to the CDC Web site. Use the search facility at the CDC site to locate pages containing information on testing for HIV. You will find that various types of immunoassay are useful for HIV testing. What are these immunoassay types Then use Google to search the Web for these types of immunoassay. What physical or chemical properties are used in the immunoassays What are the chemical principles behind these methods ... 

Our hollow cylindrical waveguide structure has also been coated with a human h-IgG antibody enzyme and used to detect a fluorescent labeled antigen Ag /OyB in a immunoassay-type reaction. 

In addition to studies where the nature of the recognition events per se has been the major issue, a number of application areas have been explored for imprinted matrices viz. (A) Chromatography, where the imprinted polymer is used as the stationary phase for separation and isolation (Chapter 20). This application is based on the fact that the imprinted polymer has a better retention for the template molecules than others. A related area of application is solid-phase extraction, where the imprinted polymer is used as a sponge to concentrate the molecule of interest (Chapter 23). (B) Immunoassay-type analyses in which molecularly imprinted polymers are used as antibody and receptor mimics (Chapter 25). (C) Catalysis where the molecularly imprinted polymers are used as enzyme mimics (Chapter 24). (D) Sensors and biosensor-like devices where the molecularly imprinted polymer is the recognition element (Chapters 26 and 27). 

Madersbacher S, Shu-Chen T, Schwarz S, Dimhofer S, Wick G, Berger P (1993) Time-resolved immunofiuorometry and other frequently used immunoassay types for follicle-stimulating hormone compared by using identical monoclonal antibodies. Clin Chem 39 1435-1439... 

Immunoassay type Label (s) Detection method Comments Reviews and examples... 

The terms competitive and sandwich of the two immunoassay types are descriptive of each assay procedure. In competitive inununoassay, an aliquot of enzyme-labeled antigen (Ag ) of known concentration is added to the sample for competitive binding to the primary antibody. Following this, the reaction surface is rinsed and the enzyme substrate (S) added. After allowing sufficient time for enzymatic conversion, the sample is analyzed for the electroactive enzyme product (P). The concentration of P bears an inverse relationship to the analyte (Ag) concentration because of the competitive binding of Ag. ... 

Wan et al. (2011) describe a CNT-based, multiplexing, electrochemical immunosensor utilizing a sandwich-immunoassay type on a disposable screen-printed carbon electrode for sensitive and simultaneous determination of PSA and interleukin 8 (IL-8), another cancer biomarker. 

As the result of high specificity and sensitivity, nucleic acid probes are in direct competition with immunoassay for the analytes of some types of clinical analytes, such as infectious disease testing. Assays are being developed, however, that combine both probe and immunoassay technology. In such hybrid probe—immunoassays, the immunoassay portion detects and amplifies the specific binding of the probe to a nucleic acid. Either the probe per se or probe labeled with a specific compound is detected by the antibody, which in turn is labeled with an enzyme or fluorophore that serves as the basis for detection. 

ImmunO lSS iy. Chemiluminescence compounds (eg, acridinium esters and sulfonamides, isoluminol), luciferases (eg, firefly, marine bacterial, Benilla and Varela luciferase), photoproteins (eg, aequorin, Benilld), and components of bioluminescence reactions have been tested as replacements for radioactive labels in both competitive and sandwich-type immunoassays. Acridinium ester labels are used extensively in routine clinical immunoassay analysis designed to detect a wide range of hormones, cancer markers, specific antibodies, specific proteins, and therapeutic dmgs. An acridinium ester label produces a flash of light when it reacts with an alkaline solution of hydrogen peroxide. The detection limit for the label is 0.5 amol. 

Enzyme Immunosensors. Enzyme immunosensors are enzyme immunoassays coupled with electrochemical sensors. These sensors (qv) require multiple steps for analyte determination, and either sandwich assays or competitive binding assays maybe used. Both of these assays use antibodies for the analyte of interest attached to a membrane on the surface of an electrochemical sensor. In the sandwich assay type, the membrane-bound antibody binds the sample antigen, which in turn binds another antibody that is enzyme-labeled. This immunosensor is then placed in a solution containing the substrate for the labeling enzyme and the rate of product formation is measured electrochemically. The rate of the reaction is proportional to the amount of bound enzyme and thus to the amount of the analyte antigen. The sandwich assay can be used only with antigens capable of binding two different antibodies simultaneously (53). 

Several types of labels have been used in immunoassays, including radioactivity, enzymes, fluorescence, luminescence and phosphorescence. Each of these labels has advantages, but the most common label for clinical and environmental analysis is the use of enzymes and colorimetric substrates. 

Sandwich ELISAs (Eigure 4) are the most common type of immunoassay used for the detection of proteins. A capture antibody is immobilized on the wells of a microplate. The solution containing the analyte is introduced and antibody-analyte... 

Once the immunoassay that meets the study objectives has been identified, sample collection begins. Proper sampling is critical in order to obtain meaningful results from any type of analytical assay. An appropriate sampling scheme will support the objective of the test. For example, a plant breeder may take a single leaf punch to determine quickly whether a specific protein has been expressed in an experimental plant. A more complex sampling regime would be used to determine the expression... 

Because the protein analyte is endogenous to the plant, it can be difficult to demonstrate the efficiency of the extraction procedure. Ideally, an alternative detection method (e.g., Western blotting) is used for comparison with the immunoassay results. Another approach to addressing extraction efficiency is to demonstrate the recovery of each type of protein analyte from each type of food fraction by exhaustive extraction, i.e., repeatedly extracting the sample until no more of the protein is detected. " ... 

However, if a class-selective assay is desirable (for multi-analyte assays), the handle should be located at or near a position that differentiates members of the class and exposes features common to the class. Using the pyrethroid example, an ideal immunogen should retain the phenoxybenzyl moiety and link the protein from the distal acid end (Figure 9). Using such an immunogen hapten, a class-specific immunoassay was developed that was highly cross-reactive with the type I pyrethroids permethrin, phenothrin, resmethrin and bioresmethrin. ... 

For pesticide residue immunoassays, matrices may include surface or groundwater, soil, sediment and plant or animal tissue or fluids. Aqueous samples may not require preparation prior to analysis, other than concentration. For other matrices, extractions or other cleanup steps are needed and these steps require the integration of the extracting solvent with the immunoassay. When solvent extraction is required, solvent effects on the assay are determined during assay optimization. Another option is to extract in the desired solvent, then conduct a solvent exchange into a more miscible solvent. Immunoassays perform best with water-miscible solvents when solvent concentrations are below 20%. Our experience has been that nearly every matrix requires a complete validation. Various soil types and even urine samples from different animals within a species may cause enough variation that validation in only a few samples is not sufficient. 

Anti-protozoa agents are utilized to treat diseases such as coccidiosis, which affects many farm animals, particularly poultry. Coccidiostats include polyether monocar-boxylic acid ionophores and other types of compounds. Polyether monocarboxylic acid ionophores include monesin, narasin, lasalocid, and salinomycin. The most common of these is salinomycin. Nonionophore coccidiostats include dimetridazole and halofunginone. Stanker et al. reviewed immunoassays available for coccidio-static agents. 

Additionally it has been our experience that mass spectrometry as a routine detection/identification technique for bacteria is not well received by microbiologists and clinicians who prefer less expensive, less complicated approaches to bacterial typing and identification, such as methods based on polymerase chain reaction (PCR) and enzyme-linked immunosorbent assays (ELISA). For that reason we have adapted our MS approach to serve as a means of biomarker discovery that feeds candidate proteins or leads into development as PCR targets or other immunoassay techniques. 

As an alternative, extremely sensitive detection can be achieved with reporter antibody probes tagged with intensely SERS-active compounds or with enzymes that react with substrates to yield SERS-active products. These methods often involve sandwich immunoassay techniques, which increase the number of required steps but offer the advantages of excellent sensitivity and the potential for label multiplexing. For example, Nie and coworkers recently reported the simultaneous detection of two types of antigens in a... 

While antibodies display similar structures - in fact, various types of IgGs are commonly used in immunoassays, antigens and proteins show tremendous variations in size, structure and charge strongly effecting the array performance. 

A similar type of biotin-dendritic multimer also was used to boost sensitivity in DNA microarray detection by 100-fold over that obtainable using traditional avidin-biotin reagent systems (Stears, 2000 Striebel et al., 2004). With this system, a polyvalent biotin dendrimer is able to bind many labeled avidin or streptavidin molecules, which may carry enzymes or fluorescent probes for assay detection. In addition, if the biotinylated dendrimer and the streptavidin detection agent is added at the same time, then at the site of a captured analyte, the biotin-dendrimer conjugates can form huge multi-dendrimer complexes wherein avidin or streptavidin detection reagents bridge between more than one dendrimer. Thus, the use of multivalent biotin-dendrimers can become universal enhancers of DNA hybridization assays or immunoassay procedures. 

---