Affinity chromatography immunoaffinity

Protein A chromatography Lectin affinity chromatography Immunoaffinity chromatography... 

The aqueous or organic extract obtained at this step of analysis may be a very dilute solution of the analyte(s) of interest. It may also contain coextractives, which, if allowed in the final extract, will increase the background noise of the detector, making it impossible to determine trace level concentrations of the analyte(s). To reduce interferences and concentrate the analyte(s), the primary sample extracts are subjected to some kind of cleanup including liquid-liquid partitioning, solid-phase extraction, matrix solid-phase dispersion, online trace enrichment, affinity chromatography, immunoaffinity chromatography, and ultrafiltration. In many instances, more than one of these procedures may be used in combination to increase extract purification. 

Affinity chromatography (immunoaffinity, metal affinity, chiral)... 

Other common impurities, such as immunoglobulins and protein A, result from the immunoaffinity purification of recombinant proteins or MAbs.16 If affinity chromatography is used to purify an antigen, then an ELISA can be used to detect contaminating levels of MAbs leached from the column. An assay for the antibody needs to detect the antibody in the presence and absence of its specific antigen. 

Affinity Chromatography of Immunoglobulins on immobiiized Antibodies (Immunoaffinity Chromatography, iAC)... 

Initially, the antibodies should be purified prior to prepare the immunoaffinity column. Precipitation with ammonium sulfate, ion-exchange chromatography, gel filtration chraoma-tography or affinity chromatography may be employed with the aim of antibody purification. Activated beads which are coated with bacterial proteins A or G may be used as the support material. Some parameters may be changed for the elution of the sample solution for example the ionic conditions of mobile phase may be changed or chaotropic buffers may be used [11]. 

For mycotoxin analyses radioimmunoassay (RIA), enzyme-linked immunosorbent assay (ELISAs) and affinity chromatography are the principal immunochemical methods in commercial application. Immunoaffinity columns or cartridges for specific mycotoxins are now being increasingly used in preliminary clean-up of extracts prior to final analysis by HPLC or GLC methods. 

Affinity chromatography techniques have shown less utility in analytical testing than in preparative separations for a variety of reasons, including cost and the difficulty of validating consistent operation as the column changes over time. Protein A affinity has been commonly used to quantitate the total antibody content of either ascites or cell culture fluids. To provide guidance in the development of a purification process, specific immunoaffinity resins are either available or can be readily prepared to quantitate the levels of unrelated protein contaminants. To rapidly determine what the active species in a mixture is, a monoclonal antibody that... 

Chapter 15 provides an in-depth coverage of various chromatographic methods, such as ion exchange, hydrophobic interaction, affinity, ligand, immunoaffinity, gel filtration, etc., that can be used for separations of antibodies by liquid chromatography. 

Further examples of separation techniques that exploit the asymmetric distribution of amino acid residues at the surface of folded proteins include metal ion affinity chromatography (HP-IMAC), ligand exchange chromatography (HP-LEC), immunoaffinity chromatography (HP-IAC), hydrophilic chromatography (HP-HILIC), and the various modes of biospecific (HP-BAC), and biomimetic (HP-BMC) chromatography. For example, the... 

The majority of reports have used electrospray ionization mass spectroscopy (ESI-MS) as an analytical detection method because of its sensitivity and the soft namre of its ionization procedure, which generally only leads to the detection of the molecular ions of the positive library members. Many separation techniques have been coupled to ESI-MS, including affinity chromatography (49), size exclusion chromatography (50, 51), gel filtration (52), affinity capillary electrophoresis (53-58), capillary isoelectric focusing (59), immunoaffinity ultrafiltration (60), and immunoaffinity extraction (61). ESI-MS has also been used alone (62) to screen a small carbohydrate library. Other examples reported alternative analytical techniques such as MALDI MS, either alone (63, 64) or in conjunction with size exclusion methods (65), or HPLC coupled with immunoaffinity deletion (66). 

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