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A new electrochemical assay method for gene expression using hela cells with a secreted alkaline phosphatase (SEAP) reporter system
Author(s) -
Şen Mustafa,
Ino Kosuke,
Shiku Hitoshi,
Matsue Tomokazu
Publication year - 2012
Publication title -
biotechnology and bioengineering
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.136
H-Index - 189
eISSN - 1097-0290
pISSN - 0006-3592
DOI - 10.1002/bit.24461
Subject(s) - hela , alkaline phosphatase , transfection , microbiology and biotechnology , cell culture , substrate (aquarium) , reporter gene , cell , chemistry , gene expression , scanning electrochemical microscopy , enzyme , phosphatase , placental alkaline phosphatase , biology , gene , biochemistry , electrochemistry , electrode , ecology , genetics
Abstract A new electrochemical assay for the detection of secreted alkaline phosphatase (SEAP) from transfectant HeLa cells is proposed using a microarray device and scanning electrochemical microscopy (SECM). The assay consists of two steps: the first is the incubation of a transfected cell in a microarray culture device covered with a substrate modified with anti‐SEAP under physiological conditions without any additives. The array device consists of a 4 × 4 array of microwells having a size of 100 µm × 100 µm (diameter × depth). The second step is SECM measurement of secreted SEAP at the antibody‐immobilized substrate. This assay ensures accuracy and intactness because the undesired influence of endogeneous ALP is eliminated and the transfected cells are incubated in a culture device under suitable conditions. We successfully detected the expression of SEAP from intact cells at the single‐cell level using this assay. The system is useful as a cell‐based gene‐expression assay. Biotechnol. Bioeng. 2012; 109:2163–2167. © 2012 Wiley Periodicals, Inc.