Allen, Stephanie

Hong, B.J.

Jung, Y.J.

Park, J.W.

Tendler, Saul

White Rose Research Online

This is a repository copy of Biomolecule Interaction Using Atomic Force Microscope.White Rose Research Online URL for this paper:https://eprints.whiterose.ac.uk/91312/Patent:Tendler, Saul (2008) Biomolecule Interaction Using Atomic Force Microscope. . eprints@whiterose.ac.ukhttps://eprints.whiterose.ac.uk/Reuse Items deposited in White Rose Research Online are protected by copyright, with all rights reserved unless indicated otherwise. They may be downloaded and/or printed for private study, or other acts as permitted by national copyright laws. The publisher or other rights holders may allow further reproduction and re-use of the full text version. This is indicated by the licence information on the White Rose Research Online record for the item. Takedown If you consider content in White Rose Research Online to be in breach of UK law, please notify us by emailing eprints@whiterose.ac.uk including the URL of the record and the reason for the withdrawal request. (12) United States Patent ParketaL US008673621B2 US 8,673,621 B2 Mar. 18, 2014 (10) Patent N0.: (45) Date of Patent: (54) (75) (73) (*) (21) (22) (65) (63) (60) (30) Aug. 12, 2005 (51) (52) BIOMOLECULE INTERACTION USING ATOMIC FORCE MICROSCOPE Inventors: J oon Won Park, Pohang (KR); Yu Jin Jung, Pusan (KR); Bong Jin Hong, Pohang (KR); Saul Tendler, Nottingham (GB); Stephanie Allen, Nottingham (GB) Assignees: POSTECH Foundation, Pohang (KR); POSCO, Pohang-shi (KR) Notice: Subject to any disclaimer, the term of this patent is extended or adjusted under 35 U.S.C. 154(b) by 975 days. Appl. N0.: 11/673,732 Filed: Feb. 12, 2007 Prior Publication Data US 2008/0113353 A1 May 15, 2008 Related US. Application Data Continuation-in-part of application No. 11/464,481, ?led on Aug. 14, 2006, noW abandoned. Provisional application No. 60/707,892, ?led on Aug. 12, 2005, provisional application No. 60/817,608, ?led on Jun. 28, 2006. Foreign Application Priority Data (WO) .............. .. PCT/KR2005/002651 Int. Cl. C12M1/00 C12M1/34 C12M3/00 C12Q 1/68 C07H 21/02 C07H 21/04 US. Cl. USPC 435/283.1; 435/6.1; 435/287.1; 435/287.2; 536/23.1; 536/24.3; 977/850; 977/863; 977/924 (2006.01) (2006.01) (2006.01) (2006.01) (2006.01) (2006.01) (58) Field of Classi?cation Search USPC ....... .. 435/6.1, 283.1, 287.1, 287.2; 536/23.1, 536/24.7; 977/850, 863, 924 See application ?le for complete search history. (56) References Cited U.S. PATENT DOCUMENTS Colton et al. .............. .. 435/6.19 Henderson et a1. Xu et al. ....................... .. 73/105 Quate et al. Park et al. ................... .. 514/489 Schlaf et a1. Cai et al. ..................... .. 427/299 Shah et al. ...................... .. 435/6 5,372,930 A * 12/1994 5,763,768 A 6/1998 5,874,668 A * 2/1999 6,436,647 B1 8/2002 2003/0199577 A1* 10/2003 2004/0009308 A1 1/2004 2004/0213910 A1* 10/2004 2005/0032060 A1* 2/2005 FOREIGN PATENT DOCUMENTS WO02/20469 * 3/2002 WO02/33412 * 4/2002 OTHER PUBLICATIONS Chen et al, Interaction between glutathione and glutathione-S transferase on dendron self-assembled controlled pore glass beads, 2004, Tetrahedron, 60, 7293-7299.* Consolandi et al, Two Ef?cient Polymeric Chemical Platforms for Oligonucleotide Microarray Preparation, 2002, Nucleosides, Nucleotides and Nucleic Acids, 21, 561-580.* * cited by examiner Primary Examiner * Narayan Bhat (74) Attorney, Agent, or Firm * Don D. Cha; Hamilton DeSanctis & Cha, LLP (57) ABSTRACT The present patent application describes a cantilever for atomic force microscopy (AFM), Which includes a cantilever body having a ?xed end and a free end, the free end having a surface region being chemically modi?ed by a dendron in Which a plurality of termini of the branched region of the dendron are bound to the surface, and a terminus of the linear region of the dendron is functionaliZed. 25 Claims, 18 Drawing Sheets US. Patent Mar. 18, 2014 Sheet 1 0f 18 FIG.1A 75 light beam position detector light source controller US 8,673,621 B2 60 US. Patent Mar. 18, 2014 Sheet 2 0f 18 US 8,673,621 B2 FIG.1B US. Patent Mar. 18, 2014 Sheet 3 0f 18 FIGZA FIG.2B AFM Tip —> The end of the AFM Tip US 8,673,621 B2 US. Patent Mar. 18, 2014 Sheet 4 0f 18 US 8,673,621 B2 FIG. 2C Pointprube® Plus supersllaripsilicon "" Tipless Cantilever Diamond coated Hard Magnetic Coating Special Developments High Aspect Ratio US. Patent Mar. 18, 2014 Sheet 5 0f 18 US 8,673,621 B2 FIG.3 FIG.4A 37 pH up ‘6 pH IINIIIIIIIQIIOIIBIII FIIIII US. Patent Fora-[pH] Mar. 18, 2014 Sheet 6 0f 18 US 8,673,621 B2 M Dakar“- lul FIG. 4C IIIIIIIIII Foro?pll] US. Patent Mar. 18, 2014 Sheet 7 0f 18 US 8,673,621 B2 FIG.5A ‘ 3? pH on. an -i a: w 1 g ... II: II .J I l I l l I l B I I I d I n I 1-.- pl] FIG. 5B Force (pN) Distance (nm)I US. Patent Mar. 18, 2014 Sheet 11 0f 18 US 8,673,621 B2 FIG. 9 US. Patent Mar. 18, 2014 Sheet 13 0f 18 US 8,673,621 B2 FIG. 13 Competition Protein —r US. Patent Mar. 18, 2014 Sheet 14 0f 18 US 8,673,621 B2 FIG. 14 20 nm W H Bil 10B 120 "D 16'] 180 210 20 Farce (pH) 90 10] 120 ND 1E1] 13D ‘29D 20 US. Patent Mar. 18, 2014 Sheet 15 0f 18 US 8,673,621 B2 FIG. 15 8O - 70 _ 6U _ 50 Force (pN) 8 S l M O l 11 %—1 ' ' I ' I ' I’!I I ' I ' I ' I ' u so 100 150 Too sou 9:10 1000 Concentration of PLC-r1 (nM) US. Patent Mar. 18, 2014 Sheet 16 0f 18 US 8,673,621 B2 FIG. 16 (a) “hw- 0-; (C) " (d) "' "' 60 p" u 54 P" g‘ u g as ‘ E I I Dlhlnl (p) nilnu (p) US. Patent Mar. 18, 2014 Sheet 17 0f 18 US 8,673,621 B2 FIG. 17 a: pH N p l a n pl m 1 1. H I Inn! pl; US. Patent Mar. 18, 2014 Sheet 18 0f 18 FIG. 18 Before Blocking S 1m pH 60 llllll Alter llocklng GIpN 'i' Injection 'E ‘g’. —"‘ ‘I. w Free a? WKYI'IVI'II )8 pm “PM in") US 8,673,621 B2 US 8,673,621 B2 1 BIOMOLECULE INTERACTION USING ATOMIC FORCE MICROSCOPE CONTINUING DATA The present application is a continuation-in-part of US. patent application Ser. No. 11/464,481, ?led Aug. 14, 2006, noW abandoned Which claims priority to US. Provisional Patent Application No. 60/707,892, ?led Aug. 12, 2005, and US. Provisional Patent Application No. 60/817,608, ?led Jun. 28, 2006, the contents of Which are incorporated by reference herein in their entirety. This application also claims priority to PCT/KR2005/002651, ?led Aug. 12, 2005, Which claims priority to US. Provisional Patent Application No. 60/601,237, ?led Aug. 12, 2004, the contents of Which are incorporated by reference herein in their entirety. BACKGROUND OF THE INVENTION (a) Field of the Invention The present invention relates generally to atomic force microscopy (AFM), a cantilever for AFM, and an apparatus and a measuring method of intermolecular interaction betWeen the biomolecules using the same. The present inven tion regards the usage of dendron coated Bio-AFM tips in measuring the interaction force betWeen biomolecules. The present invention also provides details on Bio-AFM Force Mapping of cell receptors by using surfaces With controlled meso spaces. (b) Description of the Related Art In the post-genomic era, quantitative and comprehensive studies on genome for drug discovery, as Well as disease diagnosis and prevention, are fast-groWing research and development areas. GroWth in these sectors has already pro duced a strong demand for advanced biomolecular recogni tion probes With high sensitivity and excellent speci?city (K. Wang et al., Anal Chem. 76, 5721 2004). Out of many biomolecular recognition studies, under standing the mechanical stability (or recognition property) of complementary DNA strands is crucial for a profound under standing of numerous important biological processes, such as DNA transcription, gene expression and regulation, and DNA replication. In this respect, stretching and force-induced melt ing of DNA have thus been investigated using several tech niques, such as optical tWeeZers, micro-pipette suction, and AFM (H. Clausen-Schaumann, M. SeitZ, R. Krautbauer, H. E. Gaub, Curr Opin. Chem. Biol. 4, 524, 2000; R. Merkel, Physics Reports 346, 343, 2001; G. U. Lee, L. A. Chris, R. J. Colton, Science 266, 771, 1994). As it is possible to measure speci?c interactions betWeen individual molecules at small length scales and high sensitiv ity doWn to forces of a feW piconeWtons, AFM is becoming a rapidly developing technique for probing a?inity and recog nition properties at the molecular level (R. Krautbauer, M. Rief, H. E. Gaub, Nano Lett. 3, 493, 2003). Compared With other sensitive methods for force measurements, AFM has the advantages of high force resolution and high spatial resolu tion, and is operable under physiological conditions for inves tigation of speci?c interactions in biological processes, such as electrostatic interactions (J. Wang, A. J. Bard, Anal Chem. 73, 2207, 2001), ligand-receptor binding (S. M. Rigby Singleton et al., .1. Chem. Soc., Perkin Trans. 2 1722, 2002), antigen-antibody interactions (F. SchWesinger et al., Proc. Natl. Acad. Sci. USA. 97, 9972, 2000), aptamer-protein interactions (C. Bai et al., Anal. Chem. 75, 2112, 2003), protein folding/unfolding (P. M. Williams et al., Nature 422, 446, 2003; M. S. Z. Kellermayer, S. B. Smith, H. L. GranZier, 20 25 30 35 40 45 50 55 60 65 2 C. Bustamante, Science 276, 1112, 1997), cell-cell adhesion (M. Benoit, D. Gabriel, G. Gerisch, H. E. Gaub, Nature Cell Biol. 2, 313, 2000) and DNA-DNA hybridiZation (C. W. Frank, Biophys. J. 76, 2922, 1999). Although many investigations have been performed on unbinding force measurement betWeen complementary DNA strands (H. Clausen-Schaumann, M. SeitZ, R. Krautbauer, H. E. Gaub, Curr Opin. Chem. Biol. 4, 524, 2000; R. Merkel, Physics Reports 346, 343, 2001; G. U. Lee, L. A. Chris, R. J. Colton, Science 266,771. 1994; R. Krautbauer, M. Rief, H. E. Gaub, Nano Lett. 3, 493, 2003), the recognition betWeen the DNA strands during the studies at the single molecular level is problematic. The typical immobilization approach suffered from multi-point interaction, and resolving out single molecular interactions has not been an easy task. In order to avoid the unWanted interaction, surface density Was reduced by mixing With an inactive surfactant, but the approach resulted in loW recognition ef?ciency leading to less reliable analysis. Therefore, commonly practiced surface chemistry for such immobilization such as oxide-silane and gold-thiol chemistry (T. Hugel, M. SeitZ, Macromol. Rapid. Commun. 22, 989, 2001; W. K. Zhang, X. Zhang, Prog. Polym. Sci. 28, 1271, 2003) has yet to be optimiZed to retrieve invaluable fundamental information on single DNA-DNA interaction during the force measurement With AFM. Atomic Force Microscopy has traditionally played an important role in understanding the various interaction mechanisms betWeenbiomolecules present inside organisms. Through its ability to analyZe interaction forces, its impor tance Within the ?elds of nano and biotechnology is expected to increase into the future as more studies are conducted on the molecular level. Taking advantage of this technology, many efforts have been made to investigate interactive mechanisms betWeen biomolecules on different surfaces. Unlike liquid, hoWever, observation of biological material on surfaces produce unique problems such as involuntary adsorption and steric hindrance. Among the many measures taken to counter such problems, the most popular one involve measuring monomo lecular interaction by applying complex self-assembling ?lm onto surfaces. When observing the singular interaction force betWeen tWo biomolecules WithAFM, tWo issues come to the forefront. The ?rst problem regards the difference in biomo lecular activity on a surface as opposed to Within the body. The second issue hinges around the fact that monomolecular interaction cannot be guaranteed in such a setting. Prior research has shoWn that both problems can be addressed by using self-assembling ?lm technology and meso space manipulation technology (Langmuir 2005, 21, 4257, WO 2006/016787) The technology involves observing intermo lecular interactions on the monomolecular level by control ling the spacing and number of biomolecules through limita tion of the number of functional groups on Which the molecules can be introduced. The most common problem of this method involves the unintended attraction betWeen mol ecules of the same functional group, resulting in phase sepa ration. In addition, there is no concrete evidence that the biomolecules are evenly spaced betWeen each other. The importance of biomolecular research using Bio-AFM technology is groWing rapidly. Its ability to observe noncon ductive material such as biomolecules on the nanometer level in a liquid environment that supports biological activity has made Bio-AFM an important tool in studying the structure and substructures of biological molecules. Additionally, Bio AFM enables close observation of molecular interaction by its Bio-AFM tip, onto Which a biomolecule can be loaded. Important applications include observation of interactions 

Biomolecule Interaction Using Atomic Force Microscope

https://eprints.whiterose.ac.uk/91312/1/US8673621.pdf

Biomolecule Interaction Using Atomic Force Microscope

Abstract

Similar works

Full text

Available Versions

White Rose Research Online