Immunoaffinity chromatography

Pfaunmiller, Erika L.; Bi, Cong; Zhao, Li; Vargas-Badilla, John; Zheng, Xiwei; Hage, David S.

doi:10.4155/fseb2013.14.13

Cited by 2 publications

(22 citation statements)

References 13 publications

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“…In this format, a sample containing the target analyte is applied to an affinity column in the presence of an application buffer. 11–13 During this application step, non-retained components will pass through the column, while the analyte is retained by the immobilized binding agent. The analyte is then later eluted by applying an elution buffer, followed by column regeneration.…”

Section: General Principles Of Affinity-based Methods In Hplcmentioning

confidence: 99%

“…The use of antibodies or related agents as stationary phases in chromatography is often referred to as immunoaffinity chromatography (IAC), and the use of these stationary phases in HPLC is known as high performance immunoaffinity chromatography (HPIAC). 13,14,19 Antibodies are glycoproteins that are produced by the body in response to a foreign agent, or antigen (e.g., bacteria or a virus). 13,14,19 The structure of a typical antibody, using immunoglobulin G (IgG) as an example, consists of a “Y”-shaped structure (e.g., see Figure 2).…”

Section: Affinity Methods In Hplc Using Antibodiesmentioning

confidence: 99%

“…13,14,19 Antibodies are glycoproteins that are produced by the body in response to a foreign agent, or antigen (e.g., bacteria or a virus). 13,14,19 The structure of a typical antibody, using immunoglobulin G (IgG) as an example, consists of a “Y”-shaped structure (e.g., see Figure 2). 13,14,19 The lower stem of such an antibody is known as the F c region and is highly conserved from one type of antibody to the next.…”

Section: Affinity Methods In Hplc Using Antibodiesmentioning

confidence: 99%

“…13,14,19 The structure of a typical antibody, using immunoglobulin G (IgG) as an example, consists of a “Y”-shaped structure (e.g., see Figure 2). 13,14,19 The lower stem of such an antibody is known as the F c region and is highly conserved from one type of antibody to the next. 13 The upper arms contain two identical antigen-binding sites and are known as the F ab regions; the amino acid sequences of these two regions can be highly variable between different antibodies.…”

Section: Affinity Methods In Hplc Using Antibodiesmentioning

confidence: 99%

“…13,14,19 The lower stem of such an antibody is known as the F c region and is highly conserved from one type of antibody to the next. 13 The upper arms contain two identical antigen-binding sites and are known as the F ab regions; the amino acid sequences of these two regions can be highly variable between different antibodies. 13,19 It is this variability that allows the body to produce antibodies against a wide range of targets and to produce antibodies that have strong and selective binding to these targets, with association equilibrium constants that are often in the range of 10 8 –10 12 M −1 19.…”

Section: Affinity Methods In Hplc Using Antibodiesmentioning

confidence: 99%

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High performance affinity chromatography and related separation methods for the analysis of biological and pharmaceutical agents

Zhang

Rodriguez

et al. 2018

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The last few decades have witnessed the development of many high-performance separation methods that use biologically related binding agents. The combination of HPLC with these binding agents results in a technique known as high performance affinity chromatography (HPAC). This review will discuss the general principles of HPAC and related techniques, with an emphasis on their use for the analysis of biological compounds and pharmaceutical agents. Various types of binding agents for these methods will be considered, including antibodies, immunoglobulin-binding proteins, aptamers, enzymes, lectins, transport proteins, lipids, and carbohydrates. Formats that will be discussed for these methods range from the direct detection of an analyte to indirect detection based on chromatographic immunoassays, as well as schemes based on analyte extraction or depletion, post-column detection, and multi-column systems. The use of biological agents in HPLC for chiral separations will also be considered, along with the use of HPAC as a tool to screen or study biological interactions. Various examples will be presented to illustrate these approaches and their applications in fields such as biochemistry, clinical chemistry, and pharmaceutical research.

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Section: General Principles Of Affinity-based Methods In Hplcmentioning

confidence: 99%

Section: Affinity Methods In Hplc Using Antibodiesmentioning

confidence: 99%

Section: Affinity Methods In Hplc Using Antibodiesmentioning

confidence: 99%

Section: Affinity Methods In Hplc Using Antibodiesmentioning

confidence: 99%

Section: Affinity Methods In Hplc Using Antibodiesmentioning

confidence: 99%

See 3 more Smart Citations

High performance affinity chromatography and related separation methods for the analysis of biological and pharmaceutical agents

Zhang

Rodriguez

et al. 2018

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Immunoaffinity Chromatography for Protein Purification and Analysis

Sharmeen,

Suh,

Kyei

et al. 2023

Current Protocols

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Immunoaffinity chromatography (IAC) is a type of liquid chromatography that uses immobilized antibodies or related binding agents as selective stationary phases for sample separation or analysis. The strong binding and high selectivity of antibodies have made IAC a popular tool for the purification and analysis of many chemicals and biochemicals, including proteins. The basic principles of IAC are described as related to the use of this method for protein purification and analysis. The main factors to consider in this technique are also presented under a discussion of the general strategy to follow during the development of a new IAC method. Protocols, as illustrated using human serum albumin (HSA) as a model protein, are provided for the use of IAC in several formats. This includes both the use of IAC with traditional low‐performance supports such as agarose for off‐line immunoextraction and supports used in high‐performance IAC for on‐line immunoextraction. The use of IAC for protein analysis as a flow‐based or chromatographic immunoassay is also discussed and described using HSA and a competitive binding assay format as an example. © 2023 Wiley Periodicals LLC.Basic Protocol 1: Off‐line immunoextraction by traditional immunoaffinity chromatographyBasic Protocol 2: On‐line immunoextraction by high‐performance immunoaffinity chromatographyBasic Protocol 3: Competitive binding chromatographic immunoassay

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Immunoaffinity chromatography

Cited by 2 publications

References 13 publications

High performance affinity chromatography and related separation methods for the analysis of biological and pharmaceutical agents

High performance affinity chromatography and related separation methods for the analysis of biological and pharmaceutical agents

Immunoaffinity Chromatography for Protein Purification and Analysis

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