Mechanism of Immobilized Protein A Binding to Immunoglobulin G on Nanosensor Array Surfaces

被引：56

作者：

Nelson, Justin T. ^{[1
]}

Kim, Sojin ^{[1
]}

Reuel, Nigel F. ^{[1
]}

Salem, Daniel P. ^{[1
]}

Bisker, Gili ^{[1
]}

Landry, Markita P. ^{[1
]}

Kruss, Sebastian ^{[1
]}

Barone, Paul W. ^{[1
]}

Kwak, Seonyeong ^{[1
]}

Strano, Michael S. ^{[1
]}

机构：

[1] MIT, Dept Chem Engn, Cambridge, MA 02139 USA

来源：

ANALYTICAL CHEMISTRY | 2015年 / 87卷 / 16期

基金：

美国国家科学基金会;

关键词：

WEIGHT-GAIN BIOASSAY; GROWTH-HORMONE; ANTIBODY THERAPEUTICS; ANTERIOR-PITUITARY; DIAGNOSTICS; MODEL;

D O I：

10.1021/acs.analchem.5b00843

中图分类号：

O65 [分析化学];

学科分类号：

070302 ; 081704 ;

摘要：

Protein A is often used for the purification and detection of antibodies such as immunoglobulin G (IgG) because of its quadrivalent domains that bind to the Pc region of these macromolecules. However, the kinetics and thermodynamics of the binding to many sensor surfaces have eluded mechanistic description due to complexities associated with multivalent interactions. In this work, we use a near-infrared (nIR) fluorescent single-walled carbon nanotube sensor array to obtain the kinetics of IgG binding to protein A, immobilized using a chelated Cu2+/His-tag chemistry to hydrogel dispersed sensors. A bivalent binding mechanism is able to describe the concentration dependence of the effective dissociation constant, K-D,K-eff, which varies from 100 pM to 1 mu M for IgG concentrations from 1 ng mL(-1) to 100 fig mL(-1), respectively. The mechanism is shown to describe the unusual concentration-dependent scaling demonstrated by other sensor platforms in the literature as well, and a comparison is made between resulting parameters. For comparison, we contrast IgG binding with that of human growth hormone (hGH) to its receptor (hGH-R) which displays an invariant dissociation constant at K-D = 9 mu M. These results should aid in the use of protein A and other recognition elements in a variety of sensor types.

引用

页码：8186 / 8193

页数：8

共 25 条

[1] Label-Free, Single Protein Detection on a Near-Infrared Fluorescent Single-Walled Carbon Nanotube/Protein Microarray Fabricated by Cell-Free Synthesis [J].