Chemochips [Elektronische Ressource] : development and application of analytical micro spot arrays ; a complement to universal chemical sensing / von Aniket Thete

Thete

Informations

• Publié par: technische_universitat_ilmenau
• Publié le: 01 janvier 2009
• Nombre de lectures: 29
• Langue: English
• Poids de l'ouvrage: 194 Mo

Extrait

Chemochips – Development and Application of
Analytical Micro Spot Arrays
A complement to Universal Chemical Sensing




Dissertation


zur Erlangung des akademischen Grades


doctor rerum naturalium

(Dr. rer. nat.)


Vorgelegt dem Rat der

Fakultät für Mathematik und Naturwissenschaften
der Technischen Universität Ilmenau

von M.Sc.
Aniket Thete

geboren am 27. April 1979 in Pune (India)


Betreuer: Prof. Dr. rer. nat. habil. Michael Köhler




urn:nbn:de:gbv:ilm1-2009000136


Promotionskomission


Vorsitzender: Prof. Dr. rer. nat. habil. Thomas Klar

(Institut für Physik, TU Ilmenau)



Gutachter: Prof.Dr. rer. nat. habil. Michael Köhler

(Institut für Physik, TU Ilmenau)


Prof.Dr. rer. nat. habil. Thomas Klar

(Institut für Physik, TU Ilmenau)


Prof.Dr. rer. nat. habil. Stefan Wölfl

(Ruprescht-Karls Universität, Heidelberg)


weitere Mitglieder: Dr. rer. nat. habil. Uwe Ritter

(Institut für Physik, TU Ilmenau)


Dr. rer. nat. Alexander Groß
(Institut für Physik, TU Ilmenau)













Tag des Einreichung: 21. April 2009


Tag der wissenschaftlichen Aussprache: 26. Juni 2007

























To my parents





Thesis Summary

Thesis Summary
The protein chip technology which has less developed landscape as compared to the
DNA chips, it faces the problem of universal recognition function or the strategy to
obtain a universal chip for multitude of protein samples. This is due to the chemical
complexity that protein carries as translational product of basic gene (DNA). There
exists the basic contradiction of ‘chemical recognition specificity’ against ‘universal
applicability’ in such a micro array based recognition which deals with higher chemical
complexity. It is the key issue, whether it is possible to combine different methods of
characterization and read out to make a platform by integrating specific and less specific
methods of chemically diverse analyte recognition. The issue of universal application of
chips for specific recognition of analytes has been addressed in this integration process.
The primary aim of the work is to integrate various methods of characterization and read
out, into the chip technology for liquid chemical recognition.
In order to develop the multifunctional chemochips the state of the art
characterization and read out techniques such as non-contact micro spotting,
fluorescence digital imaging and AFM techniques have been used. The spotting
technique was used to realize microarray of pure and mixed indicator functions as well
as chemical reagents for local matrix modification. Rapid fluorescence CCD imaging
techniques enables the real time recording of analyte-indicator molecule interaction by
series of images with multiple excitation-emission wavelengths. Multivariate data
analysis was applied on the pool of the data generated by the analysis of spot intensity
changes in the recorded images which gives a better idea of the analyte classification.
Along with these established methods the specially developed high resolution nano-
positioning and measurements machine equipped with AFM mode (SPM-NPMM) has
been used to characterize the micro spot array for the several millimeter range with the
resolution and precision of an AFM technique. SPM-NPMM has a large dynamic
3measurement range of 25 ×25×5 mm with the lateral resolution of 0.1 nm. The SPM-
NPMM measurements showed that the evaporation of solvent after spotting leads to the
concentric double rim formation inside the micro-spots. These characteristic double rim
features of the micro spot was found only for the binary mixed solvents with varying
polarity like water and DMF spots due to differential transport of dissolved material in
two different solvents during spot formation. High resolution SPM-NPMM
measurements showed the inhomogeneous distribution of dye nano-crystals of 2 - 5 nm

Thesis Summary

height restricted to only inner rim formed due to DMF in a double concentric rimed
spots, as the dye has less solubility in the solvent.
Using single layer chemochip arrays made up of pH and polarity sensitive dyes it
was possible to distinguish binary mixed solvents qualitatively and quantitatively. With
the help of micro array patterns, the analyte with similar content of alcohol like mixture
of 5% (v/v) of ethanol in water and beers, or other alcoholic beverages can be
distinguished qualitatively from the ethanol-water mixtures with corresponding ethanol
content. The variation in fluorescent spot patterns acted like fingerprints for complex
composed liquids like solvent mixtures and beverages. The multivariate data analysis
done on the pool of data obtained from the chemochips experiment enables to
distinguish certain classes of binary mixtures. Also, some other micro array components
like binary mixed dye spots can be classified for their response towards multitude of
liquid analytes using the PCA analysis. Analytical double layer chemochips were
prepared by splitting the functions of ‘differentiation’ in top layer and ‘indication’ in
bottom layer on the same chip. This was realized by stacking hydro-gel polymer layers
with matrix modifier using solvent casting method. The top layer was assigned with
mobility dependent differentiation by physically altering the polymer matrix using a
cross-linking agent and bottom layer functioned as indicator due to doped fluorescent
dyes. It was possible to distinguish different analytes by transition time of diffusion
through differentiation top layer by the real time recording of series of images of the
double layer arrays. The analyte molecules showed the variation in the transition time of
their transport through the top layer depending up on its cross-linking degree. This
transport of individual and mixtures of analytes through the differentiation layer can also
be detected using the same fluorescence scheme used for single layer chemochips. The
double layer chemochips also showed the analyte separation signal for a mixture of two
different analyte molecules while diffusing through the differentiation layer. This can be
attributed to the preferential interactions between analyte and the modified polymer
environment as a transport medium.
The open issue of ‘Specificity versus Universality’ faced by less developed
biochips like protein chips have been addressed through the reported work using
different types of chemochips. It was shown that different preferential and general
characterization using fluorescent micro spots was possible with single and double layer

Thesis Summary

micro arrays as a complement for the specific recognition done by the conventional spot
array chips.











Publikationen zur Dissertation / Paper based on the thesis


1. A. Thete, G. A. Groß, J. M. Köhler, „Investigation of diffusion behavior of
indicator dyes in hydro gel films for chemochip application using a microfluidic
arrangement”, VDE Micro System Technique Congress Proceedings, 2005, M22

2. A.R. Thete, G.A. Gross, J.M. Koehler, “Microfluidic arrangement with an
integrated micro-spot array for the characterization of pH and solvent polarity”
Chemical Engineering Journal 135S (2008) pages S327-S332

3. A.R. Thete, G.A. Gross, J.M. Koehler, “Differentiation of liquid analytes in gel
films by permeability-modulated double-layer chemo-chips” Analyst, 2009,
Volume 134, pages 394-400

4. A.R. Thete, T. Henkel, R. Göckeritz, M. Endlich, G.A. Gross and J.M. Koehler
“A hydrogel based fluorescent micro spot array for the characterization of liquid
analytes” Analytica Chimica Acta, Volume 633, Issue 1, 2 February 2009, Pages
81-89

5. A.R. Thete, T. Housotte, G. Jäger, N. Dorozhovets, G.A. Gross and J.M.
Koehler. “Nanotopographic characterization of spotted micro arrays on
polyvinyl alcohol films by high resolution long-range nanoprofiling”
SCANNING volume 30: 2009, pages 1–14



Propositions
• Chemochips which are motivated from the state of the art DNA chips, make use of
non-contact micro spotting, fluorescence digital imaging, and multivariate data
analysis for analyte recognition.

• The open issue of ‘Specificity versus Universality’ faced by less developed
biochips like protein chips have been addressed through the reported work of
chemochips.

• Along with the existing techniques in the field of biochips the new techniques like
high resolution long range nano-profiling using SPM-NPMM has been used for
micro spot physic