Comparison of colorimetric assays with quantitative amino acid analysis for protein quantification of Generalized Modules for Membrane Antigens (GMMA).
Comparison of colorimetric assays with quantitative amino acid analysis for protein quantification of Generalized Modules for Membrane Antigens (GMMA).
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DOI：
10.1007/s12033-014-9804-7
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发表时间：
2015-01
影响因子：
2.6
通讯作者：
Gerke, Christiane
中科院分区：
文献类型：
--
作者：
Rossi, Omar;Maggiore, Luana;Necchi, Francesca;Koeberling, Oliver;MacLennan, Calman A.;Saul, Allan;Gerke, Christiane
关键词：
Genetically induced outer membrane particles from Gram-negative bacteria, called Generalized Modules for Membrane Antigens (GMMA), are being investigated as vaccines. Rapid methods are required for estimating the protein content for in-process assays during production. Since GMMA are complex biological structures containing lipid and polysaccharide as well as protein, protein determinations are not necessarily straightforward. We compared protein quantification by Bradford, Lowry, and Non-Interfering assays using bovine serum albumin (BSA) as standard with quantitative amino acid (AA) analysis, the most accurate currently available method for protein quantification. The Lowry assay has the lowest inter- and intra-assay variation and gives the best linearity between protein amount and absorbance. In all three assays, the color yield (optical density per mass of protein) of GMMA was markedly different from that of BSA with a ratio of approximately 4 for the Bradford assay, and highly variable between different GMMA; and approximately 0.7 for the Lowry and Non-Interfering assays, highlighting the need for calibrating the standard used in the colorimetric assay against GMMA quantified by AA analysis. In terms of a combination of ease, reproducibility, and proportionality of protein measurement, and comparability between samples, the Lowry assay was superior to Bradford and Non-Interfering assays for GMMA quantification.
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DOI：
10.1074/jbc.m114.566570
发表时间：
2014-09-05
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Rossi O;Pesce I;Giannelli C;Aprea S;Caboni M;Citiulo F;Valentini S;Ferlenghi I;MacLennan CA;D'Oro U;Saul A;Gerke C
通讯作者：
Gerke C
影响因子：
3.2
作者：
FORMAL, SB;KENT, TH;LABREC, EH
通讯作者：
LABREC, EH
影响因子：
3.7
作者：
Park SB;Jang HB;Nho SW;Cha IS;Hikima J;Ohtani M;Aoki T;Jung TS
通讯作者：
Jung TS
影响因子：
2.9
作者：
KIRAZOV, LP;VENKOV, LG;KIRAZOV, EP
通讯作者：
KIRAZOV, EP
影响因子：
3.3
作者：
Forgor, AA;Leimkugel, J;Pluschke, G
通讯作者：
Pluschke, G