Highly sensitive detection of protein toxins by surface plasmon resonance with biotinylation-based inline atom transfer radical polymerization amplification.
Ultrasensitive detection of proteins is of great importance to proteomics studies. We report here a method to enhance detection sensitivity in surface plasmon resonance (SPR) spectroscopy by coupling a polymerization initiator to a biospecific interaction and inducing inline atom transfer radical polymerization (ATRP) for amplifying SPR response. Bacterial cholera toxin (CT) is chosen as the model protein that has been covalently immobilized on the surface for demonstrating the principle. The specific recognition is achieved by use of biotinylated anti-CT, which allows initiators with a biotin tag to be fixed at the protein binding site through a neutravidin bridge and triggers the localized growth of polymer brushes of poly(hydroxyl-ethyl methacrylate) (PHEMA) via an ATRP mechanism. To further enhance the signal, a second ATRP reaction is conducted that takes advantage of the hydroxyl groups of PHEMA brushes from the first step to form hyperbranched polymers onto the sensing surface. The two consecutive ATRP steps significantly improve SPR detection, allowing low amounts of CT that yield no direct measurement to be quantified with large signals. The resulting polymer film has been characterized by optical and atomic force microscopy. Ascorbic acid (AA) is employed as deoxygen reagent in the catalyst mixture that effectively suppresses oxygen interference, shortening the reaction time and making it possible for applying this ATRP approach to flow injection based SPR detection. A calibration curve of PHEMA amplification for CT detection based on surface coverage has been obtained that displays a correlation in a range from 8.23 x 10(-15) to 3.61 x 10(-12) mol/cm(2) with a limit of detection of 6.27 x 10(-15) mol/cm(2). The versatile biotin-neutravidin interaction used here should allow adaptation of ATRP enhancement to many other systems that include DNA, RNA, peptides, and carbohydrates, opening new avenues for ultrasensitive analysis of biomolecules with flow-injection assay and SPR spectroscopy.
Liu
Ying
Y
Department of Chemistry, University of California, Riverside, California 92521, USA.
Dong
Yi
Y
Jauw
Jessica
J
Linman
Matthew J
MJ
Cheng
Quan
Q
D001360:Avidin / Q000737:chemistry; D001419:Bacteria / Q000201:enzymology; D001710:Biotin / Q000737:chemistry; D019921:Biotinylation; D002772:Cholera Toxin / Q000032:analysis* / Q000737:chemistry; D004800:Enzymes, Immobilized; D057230:Limit of Detection; D018625:Microscopy, Atomic Force; D011506:Proteins / Q000032:analysis / Q000737:chemistry; D040901:Proteomics; D020349:Surface Plasmon Resonance / Q000379:methods; D013499:Surface Properties
D004800:Enzymes, Immobilized; D011506:Proteins; C093483:neutravidin; D001360:Avidin; D001710:Biotin; D002772:Cholera Toxin
Anal Chem. 2007 Jun 15;79(12):4390-400
17492838
Biosens Bioelectron. 2007 Apr 15;22(9-10):1874-80
16934448
Anal Bioanal Chem. 2009 Jan;393(1):177-86
18777170
Chem Rev. 2001 Sep;101(9):2921-90
11749397
J Virol Methods. 2006 Jun;134(1-2):164-70
16427706
J Am Chem Soc. 2004 Apr 7;126(13):4086-7
15053580
Chem Commun (Camb). 2002 Dec 7;(23):2776-7
12478741
Anal Chem. 2005 May 1;77(9):2960-5
15859616
Anal Chem. 2007 Jun 1;79(11):4233-6
17451227
Nucleic Acids Res. 2003 Sep 15;31(18):e110
12954786
Biochem Soc Trans. 2002 Aug;30(4):794-7
12196199
J Am Chem Soc. 1947 Jul;69(7):1747-53
20251412
Anal Chem. 2000 Sep 1;72(17):4193-8
10994983
Anal Biochem. 2000 Jun 1;281(2):135-43
10870828
Biomacromolecules. 2007 Dec;8(12):3922-9
18039000
Anal Chem. 2006 Feb 15;78(4):1115-21
16478102
Science. 2003 Jul 18;301(5631):352-4
12869757
Anal Chem. 2006 Jan 15;78(2):596-603
16408945
Anal Chem. 2007 Sep 1;79(17):6840-4
17685551
Anal Chem. 2007 Feb 1;79(3):1082-8
17263339
Science. 2000 Nov 17;290(5495):1315-7
11185408
Langmuir. 2006 Mar 14;22(6):2640-6
16519464
Nucleic Acids Res. 2006 Jul 05;34(11):3370-7
16822858
J Phys Chem B. 2005 Feb 24;109(7):2934-41
16851306
Biointerphases. 2006 Jun;1(2):82-92
20408620
Curr Opin Biotechnol. 2000 Feb;11(1):54-61
10679342
J Phys Chem B. 2006 Jun 8;110(22):10799-804
16771329
Anal Chem. 1998 Dec 15;70(24):5177-83
9868916
Mol Biotechnol. 2003 Mar;23(3):203-12
12665691
Biochemistry. 1999 May 4;38(18):5684-92
10231518
Anal Chem. 2006 Sep 15;78(18):6504-10
16970327
Anal Chem. 2007 Jan 15;79(2):702-9
17222040
Anal Chem. 2005 Jan 1;77(1):327-34
15623312
Analyst. 2008 Aug;133(8):975-83
18645635
Anal Chem. 2009 Feb 1;81(3):1146-53
19178341
J Am Chem Soc. 2004 Mar 24;126(11):3472-6
15025474
Anal Biochem. 2004 Oct 1;333(1):165-73
15351293
J Colloid Interface Sci. 2009 Jan 15;329(2):213-21
18950789
Anal Chem. 2005 Aug 1;77(15):4698-705
16053278
Anal Chem. 2000 Jan 1;72(1):222-6
10655657
J Am Chem Soc. 2004 Jun 2;126(21):6599-607
15161288
J Mol Graph. 1993 Jun;11(2):139-41
8347567
Biosens Bioelectron. 2008 Jun 15;23(11):1715-20
18359218
J Am Chem Soc. 2006 Nov 1;128(43):14044-6
17061884
Anal Chem. 2004 Nov 1;76(21):6173-8
15516107
Anal Chem. 2008 Nov 1;80(21):8329-33
18837517
J Mol Recognit. 2004 May-Jun;17(3):151-61
15137023
Anal Chem. 2007 Feb 15;79(4):1356-61
17297934
Opt Express. 2007 Jun 25;15(13):8163-9
19547143