Microfabrication Lab Module Georgia Tech 2011

David Gottfried Mikkel Thomas Paul Joseph Greg Book Janet Cobb-Sullivan

Lecture Outline
Introduction and biological basis for the fabricated device Cleanroom and processing basics Biological applications of microfabrication How this module will work

What this Module Will NOT Cover

Soft Lithography PDMS Microfluidics

High Density Cell Arrays

Revzin, Tompkins, and Toner, Langmuir 19, 9855 (2003) Create micro Petri dishes for study of individual surface interactions
Tissue engineering Drug screening bioassays Clinical diagnostics

Poly(ethylene glycol) (PEG) for adhesion-resistant well separations Conventional spin-coating and photolithographic techniques (top-down process) Silane treatment aids PEG covalent attachment
4

High Density Cell Arrays

Revzin, Tompkins, and Toner, Langmuir 19, 9855 (2003)

5

High Density Cell Arrays

Revzin, Tompkins, and Toner, Langmuir 19, 9855 (2003)

6

Microfabricated Cytometry Platform

Revzin et al., Lab on a Chip 5, 30 (2005) Laser scanning cytometry (LSC) as complement to flow cytometry
Direct cell visualization Small sample size Correlation of morphology and immunophenotype

Customized attachment layers (poly-L-lysine, Ab) for cell-specific interactions Batch processing of entire array Can be coupled with laser capture microdissection (LCM) for specific cell removal
7

Non-specific and Specific Attachment

Revzin et al., Lab on a Chip 5, 30 (2005)

8

Cell-Specific Attachment

Revzin et al., Lab on a Chip 5, 30 (2005)

9

Overview

Cleanroom Overview GT NRC Cleanrooms Cleanroom Etiquette & Safety Photolithography Reactive Ion Etching (RIE)

10

What is a cleanroom?
Definition: A cleanroom is an environment in which particulates, temperature, and humidity are tightly controlled to protect sensitive equipment and processes from contamination. More accurately, a cleanroom has a controlled level of contamination that is specified by the number of particles per cubic meter at a specified particle size.

11

What is a cleanroom?
ISO 14644-1 (new definition)
Class Maximum particles/m3 0.1m ISO 1 ISO 2 ISO 3 ISO 4 ISO 5 ISO 6 ISO 7 ISO 8 ISO 9 10 100 1000 10000 100000 1000000 0.2m 2 24 237 2370 23700 237000 10 102 1020 10200 102000 4 35 352 3520 35200 352000 3520000 35200000 8 83 832 8320 83200 832000 8320000 29 293 2930 29300 293000 Class 1 Class 10 Class 100 Class 1000 Class 10000 Class 100000 Room Air 0.3m 0.5m 1 m 5 m FED STD

12

Cleanroom Facilities
Pettit Microelectronics Building Opened in 1990 8500 sq. foot cleanroom space Bay and Chase design
Seven class 1000 bays Two class 10 bays

Equipment List
http://grover.mirc.gatech.edu/equi pment/

13

Cleanroom Facilities (cont.)

Marcus Nanotechnology Building Opened in 2009 15000 sq. foot cleanroom space expandable to 30000 sq. foot
10000 sq. foot inorganic cleanroom 5000 sq. foot biological cleanroom
Six BSL-1 bays Two BSL-2 bays

Labs connected by passthroughs

Ballroom style 30000 sq. feet of additional nonclean lab space

14

Georgia Tech NRC Toolset

Lithography Optical,ElectronBeam,Imprint SubstrateTreatment EtchProcessing ThermalProcessing Metallization Deposition CVD,Biomolecules Packaging Microscopy/Imaging SurfaceCharacterization ParticleCharacterization

15

Cleanroom Etiquette
Professionalism is expected from all visitors Practice clean habits
Do not touch your bare skin with your gloves Change gloves when they get dirty or are torn Don the garment in the proper manner Start at the head and work down Gloves on last Video demonstration at conclusion of lecture

16

Allowable Items
Items you cannot bring into the Cleanroom Ordinary Paper Pencils, Retractable pens Bags or purses Items you can bring into the Cleanroom Handouts on cleanroom paper Ballpoint Pens

17

Cleanroom Hazards
Potential hazards inside the cleanroom Toxic, flammable, and corrosive gases Acids and bases Solvents Electrical Sharpsrazor blades, broken wafers

18

Toxic Gas Alarms

Located on the cleanroom walls Sensors located near the gas source If the alarm goes off, evacuate the cleanroom immediately If the alarm is silenced, continue to evacuate and return only once the staff has reopened the cleanroom

19

Evacuation Procedure
Leave the cleanroom immediatelydo not de-gown Exit out nearest door this may not be the door you came in Gather at the front of the Marcus Building on Ferst Drive Remove your gown outside

20

Photolithography
Positive resist - Resist is exposed with UV light. Exposed resist becomes soluble in the designated resist developer. Negative resist Resist is exposed to UV light. Exposed resist becomes cross-linked and is resistant to the designated developer for that resist.
R. C. Jaeger, "Introduction to Microelectronic Fabrication," in Modular Series on Solid State Devices. vol. V, G. W. Neudeck and R. F. Pierret, Eds. New York: Addison-Wesley Publishing Company, 1988, p. 232.
21

Photolithography
Dosage The amount of energy required to make positive photoresist soluble in developer or negative photoresist insoluble in developer. Generally dosage is specified in photoresist datasheets in mJ/cm2. A millijoule is equal to a milliwatt-second. The power meter for the mask aligners measure power in mW/cm2.

22

Reactive Ion Etching (RIE)

Parallel plate system Top electrode showerhead RF Power Supply Plasma Density Electric field Chemical and mechanical etching

23

NanoBiotechnology Research Areas

Medical Devices Microfluidics MEMS, BioMEMS, and NEMS Biomaterials Surface Modification Biosensors Bio-metrology

24

Surface Acoustic Wave Sensors for Antigen-Antibody Interactions

D. D. Stubbs, W. D. Hunt, S. H. Lee, D. F. Doyle Georgia Institute of Technology
Non-specific target Specific target (antigen) Antibodies

Protein-A (cross-linker)

Antenna

Reflector arrays

Biolayer

The approach is to construct a vapor phase biosensor by immobilizing a monolayer of antibodies onto a surface acoustic wave sensor ((SAW) device. The binding of antigens to the antibodies studied by the change is the acoustic velocity and the oscillator frequency shifts.
25

Biosensors and Bioelectronics 17, 471 477, 2002.

Development of a Semiconductor Nanowire Sensor for the Detection of Cancer Biomarkers

Rama Ravindran, James Meindl and John McDonald Georgia Institute of Technology

The project detailed here addresses a fundamental challenge in cancer diagnosis by utilizing a silicon nanowire based sensor, fabricated using high throughput nanoimprint lithography (NIL), for the label-free detection of cancer biomarkers. Goal is early stage identification of ovarian cancer. Idea is to use silicon nanowires to detect charge of bound proteins.

Electrode

Si Wire

Electrode

26

Micro-fluidic Assay for detection of protein toxins

S. ZKuklinyek *, J. Barr *, P. Joseph, S. Aravamudhan *Center for Disease Control and Prevention (CDC) Georgia Institute of Technology

Microfluidic enzyme reactor (a) cross-sectional view, (b) overview with beads trapped in microchannel

PDMS based Microfluidic enzyme reactor connected to 10 port valve, to continuously collect products onto a HPLC column for MS/MS analysis

Protein Toxins, Botulinum and Antrax lethal factor and lethal toxin are likely agents for Bioterrorism. Screening assays must meet certain requirements: Fast, Specific, Accurate, and Sensitive.
27

Microfilters for CTC capture

S. Aravamudhan, J. Hu, P. Joseph, J. Meindl, S. Nie Emory University and Georgia Institute of Technology
Motivation CTCs in blood - significant prognostic information As few as five CTCs in about 7.5 mL of blood is clinically significant Objective Develop a MEMS parylene microfilter for a single stage selective capture of CTCs By exploiting the size difference between CTCs and normal human blood cells
Zheng et al. J Chromatogr. A 1162 (2007)

Design Detect breast (MCF-7), prostrate cancer cells (PC3) - diameters 20 22 m range. Normal red blood cells 5 9 m in diameter Microfilters 10 m thick, as 6 cm by 6 cm sheets on 4 Si wafer

28

Fabrication and Initial Results

Photoresist Silicon Parylene Silicon Silicon Photoresist Silicon Released Parylene filters
Based on the paper, Membrane microfilter for capture, electrolysis and genomic analysis of CTCs, S. Zheng, H. Lin, J.Q. Liu, M. Balic, R. Datar, R. Cote, Y. Tai, J. Chromatogr. A (2007).

Silicon

Membrane filters before capturing the CTCs

Membrane filters after capturing the CTCs

Trapped Circulating tumor cells

29

Piezo-resonator based Immunoassay for the detection of proteins expressed in cancer cells
P. Joseph, J. Meindl, F. Ayazi and S. Nie Emory University & Georgia Institute of Technology

The objective is to develop piezoelectric resonator-based Lab-on Chip (LoC) platform to detect ultra-low concentrations of cancer biomarkers Optical verification of functionalization scheme using Q-dots

30

Micromachined Resonators
Principle: Resonators are devices that naturally oscillate with amplitude at a certain frequency. When biomarkers are functionalized on resonator surface, mass loading causes decrease in resonance frequency. When resonators are made of piezoelectric material (ZnO), mechanical stress associated with the oscillation during resonance leads to a voltage differential across the device Proof-of Concept Experiments: Tumor Necrosis Factor Alpha (TNF-) and CA-125 Functionalization Scheme: SAM, EDC-NHS chemistry on Au surface

31

Micromachined Resonator Design

Bulk acoustic resonator with annexed sensing platforms Piezoelectric biosensor has two annexed sensing platforms that are patterned with a bio-affinity coating. Green squares indicate where the sensing layer will be deposited on the platform. Interaction between antibody and antigen will cause a change in mass which will, in turn, affect the resonant frequency of the resonator

32

Optical Validation
Tagged 2nd Ab

Substrate

ShortEDCincubation

LongerEDCincubation

= QD = Target Antigen

Modified Au substrate

QD

To verify that the SAM/EDC chemistry was working, biotinylated antibodies with attached QDs were used to covalently attach to the activated SAM, giving the results seen above

33

National Nanotechnology Infrastructure Network (NNIN)

Mission: Enable rapid advancements in science, engineering and technology at the nanoscale by efficient access to nanotechnology infrastructure. Approach: A network of shared open facilities distributed throughout the country that will enable the full creative abilities of the nanoscale user community to emerge.

GT NRC:

Application of nanofabrication to bioengineering and biomedicine Lead on K-12 education for the network

34

GT NRC User Support Users Statistics GT NRC NanoBio Users

Academic Albany State Univ. Clemson Univ. Emory Univ. Georgia State Univ. Georgia Tech Georgia Tech Research Institute Louisiana State Univ. Medical College of Georgia Mercer University Univ. of South Carolina Univ. of Tennessee Vanderbilt Univ. Industry Axion Biosystems Biotronics CardioMEMS Celltaxys CibaVision Greatbatch, Inc. Icon Interventional Kimberly-Clark nGimat Visigen Biotech Government Centers for Disease Control and Prevention

689 Total Users (2010) ~1/3 External

External Industry Users

External Academic and Govt Users GT Users

35

Process Flow

Clean Plasma Silane Spin coat Exposure Development

Piranha bath, rinse, dry O2 plasma 3-acryloxypropyl trichlorosilane Poly(ethylene glycol) diacrylate 2,2-dimethoxy-2-phenylacetophenone Mask aligner (365 nm) DI H20

36

Characterization
Scanning Electron Microscopy high resolution imaging

Contact Angle surface hydrophobicity/hydrophilicity

Confocal Microscopy 3D optical profiling and measurement of fabricated structures

37

Process Modifications
Use of Si/SiO2 or glass substrate Optimization of spin speed and time for thickness Optimization of UV exposure and development times Deposition of attachment layer using site-specific microcantilever rather than batch method

38

Lab Module Components

Facility Tour Leader: Paul Fabrication (Inorganic Cleanroom) Leader: Mikkel SEM Imaging (Inorganic Cleanroom) Leader: Greg Characterization and Deposition (Organic Cleanroom) Leader: David

39

Schedule
Session 1 Session 2 5 Orange TA: Philip Blue TA: Alexis Red TA: Amy Green TA: Rachel Tour Organic M i n u t e B r e a k Fab SEM Session 3 Session 4

Fab

SEM

Organic

Tour

SEM

Fab

Tour

Organic

Organic

Tour

SEM

Fab

40