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Single Cell Biosensor. Allan Fierro David Sehrt Doug Trujillo Evan Vlcek Michael Bretz. Fabrication. Flow Control. Introduction. Optical Detection Circuit. Allan Fierro. Cell Trapping. What is Flow Cytometry?. Technique for counting cells Examining cells Sorting cells.

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single cell biosensor

Single Cell Biosensor

Allan Fierro

David Sehrt

Doug Trujillo

Evan Vlcek

Michael Bretz

introduction

Fabrication

Flow Control

Introduction

Optical Detection Circuit

Allan Fierro

Cell Trapping

what is flow cytometry
What is Flow Cytometry?
  • Technique for counting cells
  • Examining cells
  • Sorting cells
example of flow data
Example of Flow Data

Analysis of a marine sample of photosynthetic picoplankton by flow cytometry showing three different populations (Prochlorococcus,Synechococcus and picoeukaryotes)

what is optofluidics
What is Optofluidics?
  • Optofluidics combines microfluidics and optics
  • Able to see the difference in light refraction

wavelength

wavelength

*diagrams courtesy of L. Shao, Ph.D.

from Ph.D. Defense Presentation

flow cytometry vs optofluidics
Flow Cytometry vs. Optofluidics

Flow Cytometry

Optofluidics

  • Flow uses scatter of light
  • Flow more expensive to run
  • Flow samples take longer to prep
  • Can’t use cells after
  • Optofluidics uses refraction of light
  • No fluorescent dye - cheaper and less prep time
  • Optofluidics can keep cells after
slide7

Microscope

Light

Cell

Infrared LED

fabrication

Fabrication

Flow Control

Fabrication

Optical Detection Circuit

David Sehrt

Cell Trapping

dep chip

DEP Chip

Images courtesy of Weina Wang

dep chip processing
DEP Chip Processing

1. 30 nm Chrome Deposition

2. 120 nm Gold Deposition

3. Spin Photoresist

4. Exposure

5. Development

6. Gold Etch

7. Chrome Etch

8. Resist Removal

pdms channel
PDMS Channel
  • PDMS- Polydimethylsiloxane
  • Suitable optical properties
  • Adhesive to glass
  • Channel mold made of a silicon substrate and SU-8 Photoresist
  • PDMS poured into mold and baked to form elastic solid
dep pdms bonding
DEP PDMS Bonding
  • Oxygen-plasma treated
  • Bonding is performed with a mask aligner
  • Minute Pressure is applied for bonding to transpire

DEP Chip

PDMS Channel

flow control

Fabrication

Flow Control

Flow Control

Optical Detection Circuit

Evan Vlcek

Cell Trapping

chip design for fluid
Two drilled holes at each end

Nanoports for each hole

200 µm wide, 25 µm deep, channel

Need flow rate of approximately 40 µm/s

Chip Design for Fluid

drilled hole

200 µm

chip

channel

flow

nanoport

*picture courtesy of L. Shao, Ph.D.

from Ph.D. Defense Presentation

nanoport assembly
Nanoport Assembly

TOP VIEW

SIDE VIEW

chip

nanotube

nanoport

drilled hole

adhesive

o ring

adhesive

o ring

from pump

“waste”

nanoport

nanotube

block diagram
Block Diagram

DB9  RS-232

Labview

program

Oriel

Controller

Actuator

Syringe

Nanotube

Chip

Yes

Yes

Set

Parameters

Pump?

On Cycle

Off Cycle

Stop?

Stop Pump

No

actual setup
Actual Setup
  • Pump set to move actuator

0.5 µm/s

  • On for 0.825 s, off for 7 s (≈10.5% duty cycle)
  • π cm2 syringe area * 0.5 * 10-4 cm/s * 0.105  1.66 * 10-5 cm3/s through channel
  • This is 1.43 liters being pumped through the channel every 24 hours!

Oriel

Controller

actuator

0.5 µm/s

RS 232

syringe

Labview

VI

nanotube

actual setup1
Actual Setup

Labview program

actuator

syringe

Oriel

controller

optical detection circuit

Fabrication

Flow Control

Optical Detection Circuit

Optical Detection Circuit

Doug Trujillo

Cell Trapping

optical detection of cells

Optical Detection of Cells

  • -Implementation-
  • Photo diode coupled via fiber optic cable from microscope to detect light modulation
  • Monitor the change of the reverse bias current from the diode through the use of a USB Data Acquisition unit
  • Digital switch to trigger RF traps

-Purpose-

Detect the presence of a cell in proximity of a trap

Interface with the pump flow controller

Provide triggering for the RF traps

optical detection of cells1
Optical Detection of Cells

-Requirements-

  • Able to detect light modulation of cells traveling 40 µm/sec
  • Output a voltage in the range of 2V - 3.8V. Any higher voltage output may damage the DAQ
  • Photodiode must be responsive to light source of 860-910 nm

Circuit Design Flow Chart

Microscope output

Photo Diode

Low pass filter

Signal to DAQ

Amplifier Buffer

optical detection of cells2
Optical Detection of Cells

-Light Source-

The light source is set up as shown in the figure. The LED is a high-intensity Infrared LED.

Lear, Kevin L., Hua Shao, Weina Wang, and Susan E. Lana. "Optofluidic Intracavity Spectroscopy of Canine Lymphoma and Lymphocytes." IEEE Explore (2007). 4 Dec. 2007.

50/125 multimode fiber

beam-splitter

micro-fluidic sample

DAQ

DAQ

Circuit

Circuit

Labview

Labview

Traps

Traps

infrared source LEDs

optical detection of cells3
Optical Detection of Cells

-Circuit Design-

~2mV Output from Diode

A1 signal to USB DAQ

optical detection
Optical Detection

Rise Time: 128 µs

Peak Voltage:

2.7 V

Figure of Time Response of Optical Detection Circuit

cell trapping

Fabrication

Flow Control

Cell Trapping

Optical Detection Circuit

Michael Bretz

Cell Trapping

automated trapping
Automated Trapping

Analog Output

Digital Output

Data Acquisition Unit

Detection

Circuit

Triggering Circuit

AC Signal

The digital signal from the Data Acquisition Unit is input into an ADG-452 (basically a digital switch) chip. This allows an AC signal to be applied to the DEP traps

DEP Traps

dielectrophoretic dep trapping
Dielectrophoretic(DEP) Trapping
  • Uses theory of electromagnetics to trap cells
  • A non-uniform electric field causes polarization within individual molecules of spheres/cells.
  • This polarization along with electric field apply a force that will move spheres to a desired location.

Electric field

electromagnetic modeling

Ground

fluid flow

+AC Voltage

Electromagnetic modeling

Picture Courtesy of Weina Wang et al. powerpoint presentation “Lab-on-a-Chip Single Particle Dielectrophoretic (DEP) Traps” 3-6-2006.

The force caused by the electric field will push the spheres into the center of the trap.

requirements
Requirements

Sphere must be traveling at 40 micron/second or less

Signal used to generate the electric field must be a time varying signal in order to produce a non-uniform electric field.

Signal used to generate the electric field must have an amplitude greater than 5 Volts peak to peak

Both of these requirements are necessary in order for the for the force caused by the electric field to overcome the force caused by the flow of water. i.e. an object in motion stays in motion unless acted upon by a net external force.

budget
Budget
  • ADG 452 Digital Switch -- $15
  • Various circuit elements -- $10
  • Hytek iUSBDAQ U120816 -- $105
  • TOTAL EXPENSES = $130
future work
Future Work
  • Improve glass chip fabrication process
  • Fine-tune optics for better optical detection
  • Incorporate spectrometer and automate data acquisition
  • Research best trap design to allow for faster flow rate
  • Explore methods to allow for cells to be analyzed faster