Fluorescence immunoassay system based on the use of a pH-sensitive phase-separating polymer.
Poly(N-isopropylacrylamide-co-methacrylic acid) [P(NIPAAm-co-MAA)], a linear water-soluble pH-sensitive phase-separating polymer, was synthesized and used as a novel separation carrier for the reactants in immunoassay. This polymer precipitates out of water below a critical pH 5.8 at 37 degrees C and redissolves when the pH of solution is above 6.2. The characteristic of this polymer makes it possible to carry out the immunochemical steps of an immunoassay in a true solution and then to quickly separate the resulting product from the reaction mixture. The above approach was applied to determination of alpha-fetoprotein with the competitive immunoassay format. Compared with traditional ELISA using the same reactants, the proposed method was much faster (the assay time decreased from 100-120 to 30 min) and showed similar sensitivity, i.e., 0.04 ng/mL. In addition, a sandwich immunoassay method for the determination of hepatitis B surface antigen was also studied, and the results showed that the pH phase-separating immunoassay could be carried out through a sandwich or a competitive method. This general technique may also be used for a wide variety of separation processes in addition to immunoassay, in which a specific component is to be isolated for analysis, recovery, or disposal.
Yang
H H
HH
Key Laboratory of Analytical Science of MOE, Xiamen University, Xiamen, 361005, People's Republic of China.
Zhu
Q Z
QZ
Chen
S
S
Li
D H
DH
Chen
X L
XL
Ding
M
M
Xu
J G
JG
D000178:Acrylamides / Q000737:chemistry; D000911:Antibodies, Monoclonal / Q000737:chemistry; D002138:Calibration; D015200:Fluoroimmunoassay / Q000379:methods*; D006514:Hepatitis B Surface Antigens / Q000097:blood*; D006863:Hydrogen-Ion Concentration; D008689:Methacrylates / Q000737:chemistry; D011108:Polymers / Q000032:analysis; D011786:Quality Control; D000509:alpha-Fetoproteins / Q000032:analysis*
D000178:Acrylamides; D000911:Antibodies, Monoclonal; D006514:Hepatitis B Surface Antigens; D008689:Methacrylates; D011108:Polymers; D000509:alpha-Fetoproteins; C008384:methacrylic acid; C067295:N-isopropylacrylamide