[10] Covalent coupling methods for inorganic support materials

Weetall, Howard H.

doi:10.1016/s0076-6879(76)44012-0

Cited by 344 publications

(122 citation statements)

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“…Binding of imprinted and nonimprinted bovine serum albumin to p-hydroxybenzoic acid in anhydrous diisopropyl ether (19). The support was first activated with 3-aminopropyltriethoxysilane, followed by covalent attachment of albumin via glutaraldehyde and reduction with NaBH4.…”

Section: Methodsmentioning

confidence: 99%

Production of abiotic receptors by molecular imprinting of proteins.

Braco

Dabulis²,

Klibanov³

1990

Proc. Natl. Acad. Sci. U.S.A.

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When a protein is dissolved in a concentrated aqueous solution of a multifunctional organic compound, freeze-dried, and washed with an anhydrous organic solvent to remove the ligand, the resultant "imprinted" protein preparation binds up to 30-fold more of the template compound in anhydrous solvents than the nonimprinted protein in the same solvent (and both proteins in water). These artificial receptors exhibit marked ligand selectivity as well as stability in anhydrous media. This phenomenon of molecular imprinting, demonstrated for several unrelated proteins and ligands, may be helpful in the development of unique bioadsorbents and, potentially, new biocatalysts.Recent years have witnessed a surge of exciting research involving the creation of abiotic receptors for small molecules (1-6). Such artificial hosts reveal the fundamental principles of structural recognition and may be used as selective adsorbents. In addition, ligand-receptor affinity may lend itself to the induction of enzyme-like activity, as illustrated by studies on cyclodextrins (7,8) and catalytic antibodies (9,10).In addition to functioning in their natural aqueous reaction media, enzymes have recently been determined to be catalytically active in anhydrous organic solvents (11). A profound feature of enzymes in an anhydrous milieu is their high conformational rigidity, manifested in such acquired characteristics as greatly enhanced thermostability (12, 13) and ligand "memory" (14). In the present study, we have used the phenomenon of drastically lowered protein flexibility in anhydrous media (compared with water) to transform common proteins into selective artificial receptors in organic solvents. MATERIALS AND METHODSBovine serum albumin (essentially fatty acid-free), bovine erythrocyte hemoglobin (type II), chicken egg white lysozyme (EC 3.2.1.17, grade I), and poly(L-lysine) (molecular mass of -60 kDa) were obtained from Sigma. p-Hydroxybenzoic acid (Aldrich) was of 99+% purity; all other chemicals used in this study were purchased from commercial suppliers and were of analytical grade or purer.All organic solvents employed in this work were of analytical grade and were dried before use by shaking with 3-A molecolar sieves (Linde). The term anhydrous herein means that no water was detected in the solvent by the Fischer titration (15)-i.e., that the water content was below -0.01%. p-Hydroxybenzoic acid and its analogs were assayed by HPLC. Ten-microliter aliquots of ligand solutions in diisopropyl ether or another organic solvent were subjected to liquid chromatography on a Waters ,uBondapak C18 column with a mobile phase consisting of 20% acetonitrile/80% aqueous phosphate buffer (pH 2.6). The flow rate was 1.5 ml/min, and the UV absorbance of the ligands was monitored at 254 nm.L-Tartaric acid and its analogs dissolved in ethyl acetate were assayed by gas chromatography following the general method of Philip and Nelson (16). To 25 gl of a ligand in ethyl acetate was added an equal volume of bis(trimethylsilyl)trifluoroacetamide....

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Section: Methodsmentioning

confidence: 99%

Production of abiotic receptors by molecular imprinting of proteins.

Braco

Dabulis²,

Klibanov³

1990

Proc. Natl. Acad. Sci. U.S.A.

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“…There are two major different ways to conjugate the antibodies onto these magnetic carriers: one is random conjugation, and the other is oriented conjugation. The most common approaches for these two different conjugation methods are (1) zero-length linker 1-(3-dimethylaminopropyl)-3-ethyl carbodiimide (CDI) (Mehta et al, 1997;Konerachá et al, 2002;Huang et al, 2003) and (2) (3-aminopropyl) triethoxysilane (APTES) (Weetall, 1976(Weetall, , 1992Shinkai et al, 1992;Zhang et al, 2002) coupled with protein A or G, respectively. Whereas CDI can link the carboxyl group of the 1°Ab onto the hydroxyl group on the surface of nano-magnetic particle (NMP) without fixed orientation, APTES is able to achieve oriented conjugation of 1°Ab by interlinking with protein A or G. Due to the high specificity of protein A and G toward the Fc fragment of immunoglobulin G (IgG), the antigen-specific sites are left free.…”

Section: Nimp Preparationmentioning

confidence: 99%

“…The protocol for oriented conjugation of 1°Ab onto nanoparticles was modified from other literature (Weetall, 1976(Weetall, , 1992Shinkai et al, 1992;Zhang et al, 2002). Briefly, 50 mg of nanoparticle was transferred into a 50-mL solution of 10% (v/v) 3-aminopropyltriethoxysilane (APTES, Sigma-Aldrich, #A3648) in anhydrous toluene and mixed by vortex and sonication bath (10 min).…”

Section: Nimp Preparationmentioning

confidence: 99%

Nano-Immunodetection and Quantification of Mycobacteria in Metalworking Fluids

Chang

Adriaens

2007

Environmental Engineering Science

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The accurate detection and enumeration of mycobacteria in metalworking fluids (MWFs) is imperative from an environmental protection and occupational health perspective. We report here on a comparison of the labeling efficiency of nano-immunomagnetic particles (NIMP) and free antibody (FAb) to detect mycobacteria in semisynthetic MWF by using both traditional visualization analysis and cluster analysisaided visualization analysis (CAAVA). The NIMP labeling method involved coating nanometer-scale magnetic particles with Protein A, and oriented conjugation of polyclonal antibodies specific to Mycobacterium spp. The FAb labeling method is modified from the traditional immunofluorescence (IF) method for more efficient detection of mycobacteria in a model MWF. The labeling efficiency of NIMP and FAb were 7.2 Ϯ 4.6 and 16.3 Ϯ 5.5%, and the specificity 85.0 Ϯ 6.1 and 88.1 Ϯ 10.5%, respectively, based on traditional visualization analysis. Based on CAAVA analysis, the labeling efficiency of NIMP and FAb increased to 12.4 Ϯ 1.6 and 20.5 Ϯ 3.9%, and the specificity to 97.8 Ϯ 3.2 and 98.5 Ϯ 2.5%, respectively. A linear relationship of FCM counts and seeded concentrations was observed over four orders of magnitude (R 2 Ͼ 0.99) in pure and ternary cultures. The results strongly support the applicability of either FAb or NIMP coupled with CAAVA and flow cytometry for rapid detection and enumeration of mycobacteria in complex matrices.

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“…A imobilização de moléculas na superfície da ponteira pode ser em monocamada auto-organizada (SAM) com grupos tióis (-SH) 56,57 sobre filme de ouro depositado ou por meio de grupos silanóis (-SiOX) 58 em substrato de silício. Outras técnicas podem ser empregadas para ativação das superfícies com propriedades químicas específicas, como a técnica sol-gel 59 e deposição de polímeros por "spin coating" 60 .…”

Section: Monitoramento De Imunoensaios: Cantilever Do Microscópio De unclassified

Microscopia de força atômica aplicada em imunoensaios

Ferreira¹,

Yamanaka²

2006

Quím. Nova

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Recebido em 4/1/05; aceito em 3/5/05; publicado na web em 24/8/05 ATOMIC FORCE MICROSCOPY APPLIED TO IMMUNOASSAYS. Affinity reactions have been used for specific detection of their complementary partners and an enormous variety of enzyme-linked immunosorbent assay (ELISA) formats are used in research and in routine serological tests. With the advent of the atomic force microscopy (AFM) technique, the immune reactions have been monitored by these devices. In the present article we focus on applications of AFM to immunoassays. After introducing the basic concepts of AFM, a brief discussion on the monitoring of the interactions between antigens and antibodies through both topographic image and biosensor systems is presented.Keywords: atomic force microscopy; antigen-antibody; immunoassay. INTRODUÇÃONos últimos anos a construção de biossensores 1,2 tem despertado considerável interesse no monitoramento de analitos nas áreas biológica, clínica 3,4 e industrial 5 , como também no monitoramento e controle de alguns poluentes ambientais 6 . O crescente emprego de biossensores deve-se à possibilidade destes atenderem algumas exigências que os métodos clássicos de análises não conseguem, sendo as características mais relevantes: simplicidade de construção, resposta rápida, redução de custo, seletividade, sensibilidade, miniaturização, precisão, facilidade de uso e pequeno volume de amostra.Um biossensor pode ser definido como um dispositivo de detecção, contendo um componente biológico ativo (enzimas, anticorpos, antígenos, células, etc.) diretamente acoplado à superfície de um transdutor, o qual converte um sinal biológico em sinal elétrico.Com base no reconhecimento do sinal de interesse pelo detector encontram-se, por ex., os imunossensores e sensores enzimáticos. Enquanto nos biossensores com enzima são monitoradas as concentrações de substratos, produtos ou mediadores, nos imunossensores 7-9 , que são dispositivos analíticos, baseados nos princípios de imunoensaios heterogêneos e nos eventos físico-químicos, são resultantes da reação de afinidade antígeno-anticorpo (Ag-Ac).No imunoensaio, o sítio combinatório do anticorpo (Ac) ou paratopo interage especificamente com porções mais superficiais (determinantes antigênicos ou epitopos) do antígeno (Ag) ou hapteno (substância não protéica de baixo peso molecular que pode se ligar a sítios específicos de combinação de anticorpos, mas não pode, por si só, iniciar uma resposta imune), formando um complexo Ag-Ac, conforme a Equação 1. Esta interação é caracterizada por uma constante de afinidade que é função das concentrações do complexo formado, do antígeno e do anticorpo livres no meio de reação. Essa interação é mantida por forças fracas, como forças iônicas, ligações de hidrogênio, forças eletrodinâmicas (van der Waals) e hidrofóbicas, garantindo o fenômeno da especificidade antígeno-anticorpo. Também podem ocorrer reações não-específicas (reações cruzadas) de outros anticorpos que competem pela ligação com o antígeno 8,10 , como, por ex., ligação com o antígeno 8,10 .Ac ...

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[10] Covalent coupling methods for inorganic support materials

Cited by 344 publications

References 6 publications

Production of abiotic receptors by molecular imprinting of proteins.

Production of abiotic receptors by molecular imprinting of proteins.

Nano-Immunodetection and Quantification of Mycobacteria in Metalworking Fluids

Microscopia de força atômica aplicada em imunoensaios

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