Ligand binding assay methods

CHROMATOGRAPHY-LIGAND-BINDING ASSAY COUPLED METHODS, IMMUNOAFFINITY SYSTEMS, AND ONLINE (FLOW-INJECTION) LIGAND-BINDING ASSAY METHODS... 

The methods now used to measure 25-OH-D are competitive protein-ligand binding assays that use either serum globulin (diluted rat serum) (27)28) or a vitamin D-deficient rat kidney... 

S. A. Cell-free ligand binding assays for nuclear receptors. Methods Enzymol. 2003, 364, 53-71. 

The fundamental parameters for bioanalytical validations include accuracy, precision, selectivity, sensitivity, reproducibility, stability of the drug in the matrix under study storage conditions, range, recovery, and response function (see Section 8.2.1). These parameters are also applicable to microbiological and ligand-binding assays. However, these assays possess some unique characteristics that should be considered during method validation, such as selectivity and quantification issues. 

As with chromatographic methods, microbiological and ligand-binding assays should be shown to be selective for the analyte. The following recommendations for dealing with two selectivity issues should be considered ... 

Note that controversies over the technical and clinical validation of immunohistochemistry have not been completely resolved. Whether or not this method should completely replace biochemical ligand-binding assays remains controversial. Despite this cautionary statement, it is true that the specificity of immunohistochemistry is theoretically valid because it is based on the use of well-characterized monoclonal antibodies raised against epitopes restricted to the ERs. 

Ballerstadt R, Schultz JS. Competitive-binding assay method based on fluorescence quenching of ligands held in close proximity by a multivalent receptor. Analytica Chimica Acta 1997, 345, 203-212. 

Table 6.1 Ge neral differences between chromatographic methods (HPLC and LC-MS/MS) and ligand-binding assays (immunoassays).

Desilva B, Smith W,Weiner R, et at. Recommendations for the bioanalytical method validation of ligand-binding assays to support pharmacokinetic assessments of macromolecules. Pharm. Res. (2003) 20 1885-1900. 

Viswanathan CT, Bansal S, Booth B, DeStefano AJ, Rose MJ, Sailstad J, Shah VP, Skelly JP, Swann PG, Weiner R (2007) Workshop/conference report — quantitative bioanalytical methods validation and implementation best practices for chromatographic and ligand binding assays. AAPS J 9 E30-E42... 

Laboratory investigations play an essential role in medicine. Laboratory results are taken into consideration in about two thirds of all medical decisions in medical systems of industrialized countries today. The vast majority of clinical chemistry analyses are based on few analytical principles including photometry, ligand binding assays and potentiometry. For these standard methods complete automation has been achieved and multi-channel, random access analyzers realize several hundred analyses per instrument and hour on a very high level of user-friendliness. Consequently, clinical chemistry is very cost efficient today typically clinical chemistry analyses contribute less than 5 % of all costs of tertiary care hospitals. 

Viswanathan CT et al (2007) Quantitative bioanalytical methods validation and implementation best practices for chromatographic and ligand binding assays. Pharm Res 24 1962-1973... 

DeSilva, B., Smith, W., Wiener, R., Kelley, M., Smolec, J., Lee, B., Khan, M., Tacey, R., Hill, H., and Celniker, A. (2003) Recommendations for the bioanalytical method validation of ligand binding assays to support pharmacokinetic assessments of macro molecules. Pharmaceutical Research, 20, 1885 1900. 

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