Immunoprecipitation (IP) of Caco-2 Cell Lysate

Immunoprecipitation is a technique in which an antigen is isolated by binding to a specific antibody attached to a sedimentable matrix. It is also used to analyze protein fractions separated by other techniques such as gel filtration or density gradient sedimentation. The source of antigen for immunoprecipitation can be unlabeled cells or tissues, metabolically or intrinsically labeled cells, or in vitro-translated proteins. This unit describes a wide range of immunoprecipitation techniques, using either suspension or adherent cells lysed by various means.Usually, plasmas transfected cells or natural cells are ideal sample for immunoprecipitation.

The Caco-2 cell line is a immortal cells of heterogeneous human epithelial colorectal adenocarcinoma cells, developed by the Sloan-Kettering Institute for Cancer Research through research conducted by Dr. Jorgen Fogh.

Although isolated from a colon (large intestine) carcinoma, when cultured under specific conditions the cells differentiated and polarized such that their phenotype, morphologically and functionally, resembles the enterocytes lining the small intestine.

Caco-2 cell line express tight junctions, microvilli, and a number of enzymes and transporters that are characteristic of such enterocytes: peptidases, esterases, P-glycoprotein, uptake transporters for amino acids, bile acids carboxylic acids, etc.The Caco-2 cells, which exhibits a well-differentiated brush border on the apical surface and tight junctions, and expresses typical small-intestinal microvillus hydrolases and nutrient transporters, has proven to be the most popularin vitro model. 

(1) to rapidly assess the cellular permeability of potential drug candidates,
(2) to elucidate pathways of drug transport (e.g., passive versus carrier mediated),
(3) to assess formulation strategies designed to enhance membrane permeability,
(4) to determine the optimal physicochemical characteristics for passive diffusion of drugs, and
(5) to assess potential toxic effects of drug candidates or formulation components on this biological barrier.