Biomarker detection via lab-on-a-chip integrated immunoaffinity approach for fluorescence and mass spectrometry readout

Forskningsoutput: AvhandlingDoktorsavhandling (sammanläggning)

Abstract

Popular Abstract in English
The human plasma proteome containing disease-correlated protein biomarkers
reflects the pathological and physiological changes relating to diseases. These
biomarkers are of interest to aid in diagnosis, prognosis and monitoring of diseases
such as cancer. Because of the interests in the disease-correlated biomarkers, plasma,
although is a complex biological sample, spanning over 10 orders of magnitude in
protein concentrations, is the most widely used clinical specimen for analysis.
Detecting these biomarkers represents biological and technical challenges in clinical
diagnostics.
Lab-on-a-chip approach could provide insights for a simplified and improved
detection platform, eliminating the laborious sample preparations compulsory in
most conventional methods. With regards to protein biomarker detection for use in
point-of-care settings, recent advancements, paved by the microfluidic technology
have been demonstrated. Microfluidic devices, manipulating fluids in minute
volumes in the range of micro to nanoliter or less, offer insights into enabling a fully
integrated, high throughput, cost-effective, rapid ‘sample to answer’, miniaturized
immunoassay systems. Microfluidic whole-blood immunoassays hold potential in
point-of-care diagnostics application, which has brought us to the exploration of
combining the microfluidic technology and antibody microarrays described in this
thesis work. At the Department of Electrical Measurements, we have earlier
developed a sandwich antibody microarray on 3D structured porous silicon surfaces.
Using this method, in this thesis, the focus in the beginning was on detecting the
prostate cancer biomarker, Prostate Specific Antigen (PSA) in plasma samples. We
have also shown a signal enhancement step utilizing europium nanoparticles to
improve the detection limit of our microarray platform for future development of a
multiplex prostate cancer biomarker detection platform. Along the line, microfluidic
technology specifically utilizing ultrasonic standing wave forces present in the acoustic field has been extensively explored at the department. Since plasma is
highly demanded in clinical diagnostics, we designed an acoustophoresis-based
microchip capable of separating plasma from undiluted whole blood. We
demonstrated the potential application of the acoustic plasmapheresis microchip by
linking it to a subsequent off line detection of PSA on our porous silicon antibody
microarray. We then moved forward with the idea of combining on-chip plasma
separation and a microarray immunoassay, in which a manifold encompassing both
platforms in an integrated manner was designed. We showed the capability of rapid
immunoaffinity-capturing of PSA directly from whole blood samples, which hold
promise of establishing a ‘whole blood sample to answer’ assay.
Another interesting lab-on-a-chip application is in the field of mass spectrometrybased
plasma proteomics which focuses on reducing the laborious sample
preparation step prior to mass spectrometry analysis. Generally, the chip-based
capillary electrophoresis separation, on-chip protein digestion and chip for direct
infusion i.e. for ESI/MS interface are among the much earlier explored applications
in the field of proteomics. In clinical proteomics, where target proteins/ peptides are
often known, mass spectrometry analysis in combination with a pre step of
immunoaffinity separation, could eliminate the need for extensive separation
methods, offering a simplified quantitative analysis path for detection of protein
biomarkers. At the Department of Electrical Measurements, we use lab-on-a-chip
integrated immunoaffinity approach in combination with mass spectrometry that
could offer insights in specifically detecting protein/peptide biomarkers from
complex biological samples. We realized the need for integrated platforms that
include reduction in sample complexity, minimize sample transfers and the
importance to be able to enrich the targeted protein of interest prior to mass
spectrometry analysis. Using our microfabricated Integrated Selective Enrichment
Target which is compatible for use in any MALDI instrument, we developed
immunoaffinity, on-target protein biomarker digestion and sample cleanup protocols
to be able to show detection of a protein biomarker, PSA on an integrated platform.
This opens potential application for use in the development of SRM analysis for
quantitative measurements of proteins. For detecting peptide from complex samples,
we developed a porous silicon array-based immunoMALDI protocol demonstrated
by the detection of Angiotensin I peptide from plasma samples.

Detaljer

Författare
  • Asilah Ahmad Tajudin
Enheter & grupper
Forskningsområden

Ämnesklassifikation (UKÄ) – OBLIGATORISK

  • Medicinteknik

Nyckelord

Originalspråkengelska
KvalifikationDoktor
Tilldelande institution
Handledare/Biträdande handledare
Tilldelningsdatum2011 dec 16
Tryckta ISBN978-91-7473-217-7
StatusPublished - 2011