The Hong Kong University of Science and Technology Final Year Project presentation 2007 Project supervisor: Dr. Andrew Poon Department of Electronic and Computer Engineering Wong Ka Ki Chris, ee_wkkaf, 04067147 Wong Tsz Ong Mark, ee_wto, 04319837 Wong Tsz Ming Keith, ee_wtm, 04129115
Introduction
A B C Bottle A Bottle B Bottle C C A B? Bottle C Bottle A Bottle B
Project Objective Preliminary testing and sorting of droplets based on light scattering principles. Fast and non-intrusive such that the nature of sample would not be affected.
Droplet Generator. Pinhole STEP 1 vibrate Droplet filter Speaker Function Generator Gated Red Laser 60º CCD 1 555 timer E CCD 2 STEP 3 42º Droplet Objective Lens To PC (Interferometic Particle Imaging ) STEP 2 To PC (Rainbow Refractometry) Air Puffer PC Data input from CCDs Glass Plate CCD 1 CCD 2 STEP 5 STEP 4 Wanted droplet Unwanted droplet Move using step motor Motor
System Flow Five Steps Approach [Step 1]: Droplet is produced by droplet generator. [Step 2]: Droplet size is determined by CCD1 (Interferometric Particle Imaging) [Step 3]: Droplet refractive index is determined by CCD2 (Rainbow Refractometry) [Step 4]: Computer analysis. [Step 5]: Unwanted droplets are sorted by air puffer
Project Advantages & Applications
Project Advantages Fast-testing Preliminary Result Non-intrusive Non Bio-Chemical
Applications Pharmaceutical Industries Medical Testing Bacteria Medicine Composition Industrial workplace Pigment
Droplet Generator. Pinhole Function Generator Upstream vibrate Speaker Droplet filter Gated Red Laser 60º CCD 1 555 timer CCD 2 42º Droplet Objective Lens To PC (Interferometric Particle Imaging) Mid-stream Wanted droplet Unwanted droplet To PC (Rainbow Refractometry) Glass Plate Air Puffer Syringe Knob PC Data input from CCDs Down-stream Move using step motor Motor
Upstream
Components [Droplet Generator] -Produces a stream of droplet samples -Manual control of production rate [Droplet Filter] -Falling path control
Droplet Generator: To start [Primary ideas]: (1) Syringe? (2) Burette? (3) Dropper? (4) Spraying bottle?
Droplet Generator: Structure [Building unit]: (1) Loudspeaker (2) Syringe (3) Plastic tape (4) Optical fiber (5) Function generator [Where does this idea come from?] [Building steps]
Droplet Generator: Performance Droplet production rate can be controlled by the function generator Droplet size is approximately 270µm in diameter [Problem]: - Falling path deviation - Plastic tape depreciation
Droplet Filter [Any water-resistive materials / tools]: -Laser aperture control -Screw -Plastic taped surface with a tiny hole in the middle Video number: P1010061
Mid-stream
Components [Gated laser source] -Hitting the sample droplet to gives out scattering patterns -As a light source for image capturing [CCD cameras] -Located at 42 0 and 60 0 respectively -Capture light scattering patterns
Why Gated laser source?? Blurred images Overlapping of images even from a single sample droplet Limitation of the shutter s speed of the CCD camera
Gated laser source Stroboscopic Investigation technique Overlapping of images is resolved Pulse duration = 0.4µs
Image Capturing Two light scattering techniques were used: Droplet size measurement Interferometric Particle Imaging (IPI) Droplet Refractive Index determination Rainbow Refractometry What is Light Scattering?
Light Scattering from Droplet Interaction of a laser beam with droplets is considered Incident plane wave k The laser is operated in the TEM00 mode creating a beam with a profile of Gaussian like y φ r x Detector When the droplet is comparably small to the diameter of the laser source, the Gaussian beam can be considered to be a plane wave The interaction between the laser beam and the droplet results in light scattering in all directions around the droplet Droplet θ z Scattering plane Polarization plane The scattered light can be captured by a receiver, for example, a CCD camera which is placed on the scattered plane The region with (0º θ 90º) is called forward scattering region, whereas the remaining hemisphere is called backward scattering region. The intensity distribution can be calculated with Lorenz-Mie Theory
Light Scattering from Droplet Forward Scattering Region Backward Scattering Region 2 nd rainbow 1 st rainbow Light Source Related intensity versus scattering angle Plotted by MiePlot. (n =1.334, λ =650nm, droplet diameter = 20µm) In the forward light scattering region, interference patterns with regular fringe spacing can be observed and part of this scattered light can be used to determine the Falling droplet droplet size. (Interferometric Particle Imaging) In the backward light scattering region, rainbow fringes are observed and they are droplet content dependent, for example texture and refractive index. (Rainbow Refractometry) Red Laser
Theories of Light scattering Why such intensity distribution comes out? If the droplet size is much smaller than the wavelength of the laser source (d << λ) Raleigh theory will be applied. If the droplet size a much larger than the wavelength of the laser source (d >> λ) geometrical optics will be applied However, geometrical optics does not provide an accurate result in scattering angles Lorenz-Mie theory. It works well for all droplets sizes and scattering angles. This theory came out from Lorenz (1890) and Mie (1908). It is a wave theory that is a solution of Maxwell s equations for Electromagnetic waves. Geometrical optics has deficits but it still helps to understand the light scattering phenomena Incident Light P = 0, Reflection droplet θ P = 1,1 st Refraction By Snell s law The 1st refraction is on the forward scattering region. The interference of these refracted lights together with the reflected light results in the regular interference fringes pattern While the scattered angle of different light rays from the 2nd refraction attains maximum at a certain angles, which is called rainbow angle P = 2, 2 nd Refraction
Rainbow Refractometry Refractive index is an important parameter for one to obtain additional information from a droplet Several laser-based techniques have been investigated. E.g. fluorescence and intensity-based light scattering Fluorescence: Additives are added and these pollute the droplet sample Intensity based light scattering: The intensity distribution of the scattered light is used to determine the droplet refractive index. E.g. Rainbow Refractometry The interference between internally reflected rays will induce a low frequency structure called Airy fringes. The interferences between internally and externally reflected rays will cause a high frequency structure called ripple structure superimposed on the Airy fringes Airy Theory Lorenz Mie theory Diameter: 200µm Refractive Index: 1.33 Wavelength: 650 nm Lorenz-Mie theory, we can compute the rainbow scattered by a single droplet. This helps us to determine the refractive index of the droplet
Rainbow Refractometry How the refractive index is determined? n = 1.33 n = 1.34 n = 1.35 The refractive index is determined from the absolute angular position of the interference pattern, the pattern shifts uniformly for about 1.5º when the refractive index changes by 0.1 Another method such as the relative intensity of the first few peak of the fringes of different are measured to obtain the refractive index. However, the pattern is size dependent, droplet size should be known Interferometric Particle Imaging (IPI)
Interferometric Particle Imaging (IPI) Technique used to measure droplet size Top View Non Focal Plane Focal plane 1 st refraction z Focused Image Droplet θ α da Reflection Aperture Red Laser Beam Lens Image Plane d = 2 λn msin( θ / 2) 1 α (cos( θ / 2) + m 2 ) 2mcos( θ / 2) + 1 N is the number of fringes, λ is the source wavelength, m is the refractive index of the liquid droplet, θ and α are the scattering and collecting angles, respectively. α is equivalent to the product of fringe number N and angular inter-spacing Δδ
Interferometric Particle Imaging (IPI) The dependence of the size measurement on the refractive index reduces to a minimum as a scattering angle of 60. In the other word, the diameter measurement is less dependent on the refractive index of the droplet. By measuring the number of fringes captured by a CCD camera non focused plane, droplet diameter can be obtained by the equation.
Down-stream
Sorting System Electrostatic Sorting Air Puffing
Electrostatic Wanted Cell Unwanted Cell - - - - - - - - - - - + + + + + + + + + + +
Electrostatic Implementation Difficulties Charge Injection High Voltage (Hazard)
Air Puffer Advantages: Relatively Safe Syringe Easy to Control Plastic membrane Needle
Air Puffer Real Time On Substrate Glass Plate Syringe Wanted droplet Unwanted droplet
Real Time Puffer Triggering signal Plastic membrane MP3 layer Amplifier magnet Loud Speaker 3W Syringe Needle
On Substrate Puffer Knob Plastic membrane nail Syringe Needle Motor
Result
Droplet size measurement Water, 3% glucose, 10 % glucose having different refractive index were measured. Number of interference fringes were counted by ImageJ. Using the equation from IPI, we can obtain the droplet size. Scattering angle of around 60 was used to obtain a lesser dependence of the refractive index on the droplet size measurement. In this experiment, λ = 650nm. α = 0.07995 rad and θ = 60º 10 samples were used for the calculation in each set of data. Water Refractive index is 1.33 at room temperature. Average number of fringes = 30.6 Droplet diameter = 270.3 μm 42.0
Droplet size measurement 3% Glucose Solution Refractive index is 1.3375 at room temperature. Average number of fringes = 33.1 Droplet diameter = 270.35 μm 10% Glucose Solution Refractive index is 1.355 at room temperature. Average number of fringes = 33.8 Droplet diameter = 272.22 μm
Droplet size measurement The average diameter of a falling droplet calculated by the IPI equation is 270.1μm, which is close to estimated droplet diameter from the photo.
Rainbow Refractometry P = 0, Reflection Droplet Incident Light P = 1,1 st Refraction θ P = 2, 2 nd Refraction (Rainbow Region) Backscattering
Rainbow Refractometry HeNe gas Laser Wavelength ~ 632.8nm Optical power ~ 20mW
Rainbow Refractometry
Rainbow Refractometry Droplet Intensity Measurement Relative Intensity 80 70 60 50 40 30 20 10 0 0 200 400 600 800 Pixel (Distance)
Rainbow Refractometry Water (sample): Water 100 80 Intensity 60 40 20 0-20 0 200 400 600 800 Pixel (Distance) Contrast Ratio = (55/96) X 100% = 56%
Rainbow Refractometry 10% Glucose Solution (sample): 10% Glucose Solution Intensity 120 100 80 60 40 20 0-20 0 200 400 600 800 Pixel (Distance) Contrast Ratio = (63/104) X 100% = 60.5%
Rainbow Refractometry 30% Glucose Solution (sample): 30% Glucose Solution 120 100 80 Intensity 60 40 20 0-20 0 200 400 600 800 Pixel (Distance) Contrast Ratio = (73/110) X 100% = 66.4%
Rainbow Refractometry Summary Table: Contrast Ratio Water 10% Glucose 30% Glucose 47.5 0 0 0 50 2 0 0 52.5 1 2 1 55 3 1 2 57.5 6 2 2 60 7 3 4 62.5 2 8 2 65 3 4 10 67.5 1 3 5 70 1 1 1 72.5 0 2 2 75 2 1 0 77.5 0 2 0 80 0 0 1 82.5 0 0 30 captured photos for each solution 0
Rainbow Refractometry Water 10 % Glucose 30% Glucose
Rainbow Refractometry Frequency 12 Comparison Between Contrast Ratio Verus Differnent Aqeous Solution 10 8 6 Water 10% Glucose Solution 30% Glucose Solution 4 2 0 45 50 55 60 65 70 75 80 85 Contrast Ratio
Conclusions
Limitations Preliminary test Affected by air flow
Limitations
Eccentricity Limitations
Droplets Shape Limitations
Limitations Sensitivity ( n???) Solution Water 10% Glucose 30% Glucose Refractive Index 1.33?? Concentration = Mass of Glucose (g) Water Volume (ml) X 100%
Limitations Example. Slope = 2.5µ n/mg/dl g/ml mg/dl (10% Glucose) 1g/10mL 10000 mg/dl n = 0.025
Limitations Solution Refractive Index Water 1.33 10% Glucose 1.355 30% Glucose 1.405 Sensitivity = 0.03
Improvement Smaller droplet can be generated by ink-jet printer Scattering angle detection can be used as an additional information to the change in refractive index The glass plate can be replaced by a Micro-fluidic cells that droplets can be further investigated bio-medically
Example on the usage of micro-fluidic cells Data from CCD Data from Micro-fluidic cell Microscope Air Puffer Output Signal Puff of air hydraulic pull Unwanted Droplet Wanted Droplet Micro-fluidic cells Light ray Diode
Contributions & Further work
Acknowledgements & Final Words