navin aptamer

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Presenting by: Course name: Microbial GeneticsNaveen kumara.A.C Course no:DM-612

1st

year M Tech(DM)

Aptamer-linked Gold nanoporticle forcolorimetric sensing of analytes


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y Aptamers are single-stranded DNA or RNA molecules thatcan bind target molecules with high affinity and specificity.

y Aptamers can be obtained by a combinatorial biologymethod called systematic evolution of ligands byexponential enrichment (SELEX).

y The conformation of an aptamer usually changes upon

binding to its target analyte.

y This property has been used in a wide variety of sensingapplications, including detection based on fluorescenceintensity, polarization, energy transfer, electrochemistry or colour change.

Introduction


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y C olorimetric sensors are particularly important

because they minimize or eliminate the necessity of using expensive and complicated instruments.

y The many colorimetric sensing strategies, metallicnanoparticle-based detection is desirable because of the high extinction coefficients and strong distance-dependent optical properties of the nanoparticles.

y Aptamer-linked gold nanoparticle purple aggregatesthat undergo fast disassembly into red dispersednanoparticles upon binding of target analytes.


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y

The adenosine-sensitive nanoparticle aggregatesconsist of three componentsTwo DNA-functionalized nanoparticles(particles 1and 2)

A linker DNA (Linker Ade ).DNA sequences and modifications.


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y The DNA for particle 1 is attached to a nanoparticle at

its 3 end.

y The linker DNA is designed so that the 5 end, which iscomplementary to the DNA attached to particle 1, is

separated from the adenosine aptamer at its 3 end by apentanucleotide sequence.

y The DNA for particle 2 is attached to a nanoparticle at

its 5 end and is complementary to the pentanucleotideand to the first portion of the adenosine aptamer.

y In the absence of adenosine or in the presence of other molecules such as other nucleosides, the aggregates


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y In the presence of adenosine,however, the aptamer DNA(green) switches its structure and binds two adenosine

molecules.

y As a result, only the pentanucleotide ( in gray) in the linker DNA is left to bind particle 2.

y The five DNA base pairs are not strong enough to holdparticle 2 at room temperature, leading to its dissociationand resulting in red individual gold nanoparticles.

y The most important element in designing such stimuli-

responsive aggregates is the number of base pairs betweenthe linker DNA and the DNA on particle 2, and the number of nucleotides from the aptamer sequence that are involvedin binding to particle 2.

y

The binding should be strong enough to hold thenanoparticles together but still allow the aptamer to bind


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Colorimetric detection of adenosine with aptamer-assembled nanoparticle aggregates.

a.UV-visible spectra of dispersed (red)and aggregated (blue) goldnanoparticles.c. Kinetics of color change of adenosine

aptamer-assembled aggregates in thepresence of 1 mM nucleosides. Inset:photograph of the four samples withdesignated nucleoside added.d.Kinetics of color change of theaggregates with varying adenosineconcentrations.


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y O ne major challenge in analytical chemistry is multiplexsensing of a number of analytes with each analytedisplaying a different signal.

y Quantum dots(QDs) that emit at 525 and 585 nm areused to encode aptamer-linked nanostructures sensitiveto adenosine and cocaine, respectively.

y In addition to quantum dots, the nanostructures alsocontain gold nanoparticles that serve as quenchers.

y Addition of target analytes disassembles thenanostructures and results in increased emission from

quantum dots. Simultaneous colorimetric and fluorescent


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(A) Gold nanoparticles 1, 2, and quantum dot Q1were assembled by an adenosine aptamer DNA,resulting in quenched QD emission. Addition of adenosine disassembled the aggregates with

increased QD emission observed. DNAsequences and nanoparticle linkages andmodifications for the adenosine sensor

(B) and for the cocaine sensor

(A) For the cocaine sensor

QD -encoded aptamer-linked nanostructuresfor multiplex detection.

(D) Schematic of the overall structure of the QDs used inthis work (adapted from Invitrogen). In addition tothe luminescent QD core, there is a semiconductor shell,a polymer coating, and a streptavidin layer. These layers

increased the distance between the core and the goldnanoparticle quenchers, resulting in reduced quenching

(C ) Q1 and Q2 were coated with streptavidin (denoted aspink crosses) for biotinylated DNA conjugation. Q1 emitsat 525 nm and Q2 emits at 585 nm.


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y Aptamer can be selected against difficult targethaptens, such as toxins or prions.

y Aptamers can thus be considered as a validalternative to antibodies or other bio-mimetic

receptors, for the development of biosensors andother analytical methods.

y Simultaneous Detection of Multiple Analytes acts as aelectronic nose.

y These are widely used in diagnostics,therapeutics(Macugen, Archemix ).

y Aptamers can be used for both basic research andclinical purposes as macromolecular drugs.

y These are used in food industry for the detection of

App lications


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y Aptamers can be selected against essentiallyany target molecule of choice.

y The multiplex detection system should beuseful for sensing of very complex systemswith multiple target analytes.

y This method is widely accepted in in-vivodiagnostic application.

Conclusion


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