Download an integrated microfluidic fish chip: enumerating chromosomes on a

AN INTEGRATED MICROFLUIDIC FISH CHIP:
ENUMERATING CHROMOSOMES ON
A CELL-BY-CELL BASIS IN LESS THAN 1 HOUR
Vincent J. Sieben1, Carina S. Debes-Marun2, Linda M. Pilarski2 and
Christopher J. Backhouse1
1
2
Department of Electrical and Computer Engineering, University of Alberta,
Department of Oncology and Cross Cancer Institute, University of Alberta,
Edmonton, Canada
ABSTRACT
Conventional fluorescence in-situ hybridization (FISH) techniques provide a
sensitive diagnostic tool for the detection of chromosomal alterations on cell-by-cell
basis; however, the cost-per-test in terms of reagent and highly qualified labour has
prevented its wide-spread utilization in clinical settings. We demonstrate a microfluidic chip that integrates the sample preparation and reagent processing required to
perform the FISH protocol, thereby minimizing the need for human intervention.
Our microfluidic chip has lowered the reagent cost-per-test by 20-fold, decreased the
labour time by 10-fold, and substantially reduced the support equipment needed. We
believe this cost-effective platform will make sensitive FISH techniques more clinically accessible and can aid in disease management for many cancers.
KEYWORDS: DNA Hybridization, Thin-Film Heater, Bus-Valves, Integration
INTRODUCTION
For many cancers, the lack of a cost-effective and informative diagnostic method
has compromised the quality of life for patients [1,2]. Microfluidic technologies can
reduce the cost-per-test, both in-terms of reagent consumption and technician time,
making conventional assays more affordable for large-scale clinical usage. Previously we performed the FISH protocol on a microfluidic chip [1]. The first microchip iteration allowed us to reduce the cost-per-test by 10-fold by physically confining the expensive probe to a region directly over the cells. Still, the chip required a
highly trained operator to manually deliver the reagents in a stepwise manner to
complete the FISH protocol on-chip.
INTEGRATED SYSTEM
For this paper, we demonstrate a microfluidic platform that integrates and automates most of the sample preparation required to perform the FISH protocol, thereby
minimizing the need for human intervention and lowering the labour cost-per-test.
The microchip shown in Figure 1 incorporates a reagent multiplexer, a FISH chamber with an integrated thin-film heater, and a peristaltic pump. The reagent multiplexer was implemented using bus valves that have minimal dead volume, necessary
for avoiding reagent cross-contamination during the reagent processing steps. The
thin-film heater was based on previously published work and allowed on-chip thermal control [3].
Twelfth International Conference on Miniaturized Systems for Chemistry and Life Sciences
October 12 - 16, 2008, San Diego, California, USA
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Figure 1. (A) Combined mask layouts and dimensions of the integrated FISH microchip, which was employed for automating the protocol aspects of FISH. The chip
integrates a reagent multiplexer, thin-film heater, and peristaltic pump. (B) Photograph of integrated FISH chip beside a coin to illustrate the small physical size.
By integrating the FISH protocol on-chip, we have also minimized the support
equipment required; the desktop-sized system shown in Figure 2. The system was
created for less than 1000 USD, much as in reference [4], and allows a user to load
the required reagents, press start on a laptop, and perform other laboratory tasks
while the FISH protocol is performed on-chip.
Figure 2. Photographs of the support infrastructure and electronics (bottom right)
required to run the integrated FISH chip. The system provides step-wise programmatic control for 14 valves & 1 on-chip heater for fluid & temperature automation.
RESULTS AND DISCUSSION
For the first time, the entire FISH protocol was performed in an automated manner on a microfluidic device. The result is an integrated genetic test for detecting
chromosomal abnormalities in individual cells in less than 1 hour. The presented
microfluidic chip has lowered the reagent cost-per-test by 20-fold and decreased the
labour time required of a technician by 10-fold. Typically, the protocol requires
hours to days of technician time; whereas the integrated microchip requires only 5
minutes for loading reagents. As an example application for this development, we
Twelfth International Conference on Miniaturized Systems for Chemistry and Life Sciences
October 12 - 16, 2008, San Diego, California, USA
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detect the number of X and Y chromosomes per cell in several patient samples
shown in Figure 3; useful for identifying aneuploidy (an abnormal number of chromosomes per cell) that is often present in many cancers. However, a variety of
FISH probes may be purchased to detect genetic changes as subtle as 50 kbp.
(A)
(B)
Figure 3. (A) Fluorescence images acquired after the automated FISH protocol was
performed on the integrated microchip. The cell line was RMPI 8226 and two X
chromosomes (green dots) were detected per cell. (B) A male patient sample; one X
(green dot) and one Y (red dot) chromosome were detected per cell.
CONCLUSIONS
This technology is a significant step toward rapid, affordable, and automated genetic screening to recognize aneuploidy and/or specific chromosomes abnormalities
that are associated with a number of illnesses [2]. Our future work will demonstrate
a microfluidic chip capable of starting with a raw sample such as blood, and perform
all the necessary protocol steps to yield clinically relevant genetic information with
minimal human intervention.
REFERENCES
[1] V.J. Sieben et al., “FISH and chips: chromosomal analysis on microfluidic platforms,” IET Nanobiotechnology, 1 (3), p. 27 (2007).
[2] D.S. Lee et al., “Application of high throughput cell array technology to FISH:
Investigation of the role of deletion of p16 gene in leukemias,” Journal of Biotechnology, 127, p.355 (2007).
[3] V.N. Hoang et al., “Dynamic temperature measurement in microfluidic devices
using thermochromic liquid crystals,” Lab on a chip, 8, p.484 (2008).
[4] G.V. Kaigala et al., “An inexpensive and portable microchip-based platform for
integrated RT-PCR and CE,” Analyst, 133 (3), p.331 (2008).
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