The saturation prevents the oscillation of the internal magnetization of the beads and no longer contributes to the inductance change of the frequency shift sensor. 62 2.44 Synthesis and characterization of the SPION-CpG conjugates. tion of the synthesis of SPION-CpG conjugates.
Point-of-use Diagnostics
- Immunochemistry
- Molecular Diagnostics
- Pathogenomics
- Pharmacogenomics
- Genetic Testing
- Blood Donor Screening
- Point-of-Care Diagnostics
- Self-monitoring Blood Glucose and the Point-of-Use Diagnostic
- A POU Diagnostic for Protein and Nucleic Acid Detection
First, we present the landscape of an important segment of medical diagnostics – in vitro diagnostics (IVD). The amount of signal present from labeled, bound antigen or labeled, unbound antigen can be anticorrelated with the concentration of the target analyte in solution.
Magnetic Particle Trafficking for Brain Tumor Therapy
- Brain and Central Nervous System Tumors
- Malignant Glioma Treatment Options
- Magnetically Trafficking Macrophages for Guided Immunotherapy
- Prototyping a Wearable, Dynamically Programmable Magnetic Field (DPMF)
After introduction and endocytosis of CpG NPs, the macrophages would be injected directly to the tumor site. A magnetic field profile would retain particles near the tumor site while the body's immune system recruits leukocytes to the NPs.
Contributions
Organization
Introduction
In the DNA and antigen assays, the presence of biological targets leads to the accumulation of magnetic beads over the sensor. To demonstrate the viability of the sensor for biologically relevant assays, we have developed and demonstrated two representative practical assays that detect protein and nucleic acid targets [ 70 ].
Handheld, Portable Cartridge Reader
The capture DNA strand, which is complementary to the portion of the oligo strand, is attached to the PLL surface by electrostatic adsorption. The probe DNA strand is complementary to another part of the target strand and attaches to the bead via a streptavidin/biotin linker.
Easy-to-use Open Well Cartridge
The sensor array is electrically connected to the carrier and a pit is formed above the carrier. Finally, the polypropylene housing is epoxy bonded to the chip holder to form an open recess.
Magnetic Freezing
It consists of a chip carrier that is used to enable plug-and-play functionality within the reader. In a paramagnetic state, the magnetizations of the beads (Mbead) are allowed to oscillate up and down.
Surface Chemistry of Integrated Circuits (IC) for Biological Samples
For antibody attachment, an epoxy surface was prepared to allow covalent attachment to capture antibodies. The chip was then dried with nitrogen and baked at 110 °C for 15 min to covalently attach the silane groups to the glass substrate [50].
Probe Printing
Thus, the pen was regularly cleaned before printing by sonication in 50mM KOH from 10 min to 1 h [79]. After antibodies were printed on the sensors, the chip was left in the desiccator to dry overnight.
DNA Protocol
After DNA incubation, the well was washed twice each in DNA Stringent Wash Buffer 1 and 2 (Arrayit, Sunnyvale, CA) for 2.5 min each. Finally, the well was washed 3 more times with binding and wash buffer, then washed 3 times with nanopure water, and then drained.
Immunoassay Protocol
A total of 100 µl of protein solution was added to the well and incubated for 2 hours at room temperature with shaking at 125rpm. IFN-γ detection antibody (200X dilution of stock solution in assay diluent) was added to the well and incubated for 1 hour at room temperature with shaking at 125rpm. The bead solution was then added to the well and incubated for 20 minutes without agitation.
Finally, the well was rinsed 5 times with water to prevent crystallization of the remaining salt solution on the chip.
Measurement and Processing
Then the slide was rinsed 5 times with 400 µl of wash buffer (0.05% Tween-20 in PBS) to remove the blocking solution. A total of 5 µl 1µm diameter Dynabeads MyOne Streptavidin C1-coated magnetic beads were washed 3 times with PBS to remove preservatives from the bead solution [81]. After incubation, 5µl of 25% glutaraldehyde solution (Sigma-Aldrich, St. Louis, MO) was added to the well for fixation [ 82 ].
The difference between the baseline measurement and the endpoint measurement is used to derive the number of beads across the sensor.
DNA Sandwich Assay
One measurement cycle determines the basic frequency in the presence of a small rare earth magnet. At the detection limit, 100 pM, the frequency shift of the target sensor is more than twice that of a non-complementary NC sensor. At the detection limit, 100 pM, the frequency shift of the target sensor is more than twice that of a non-complementary NC sensor.
Data Processing
This is achieved by first measuring the sensor at the end of the test using the previously described frequency shift technique as shown in the figure. The number of balls above the sensor was counted (Figure 2.10) and compared with the measured signal. Areas with a printed capture strand would cause the beads to bond to the sensor surface as shown in Fig.
The detection limit for the sensor was 100 pM, with the frequency shift being more than twice the background level.
Immunoassay detection of Tuberculosis biomarker IFN-γ
Reference sensors and sensors printed with the NC capture string will exhibit some fluctuations in frequency shift measurements due to variations in background bead bonding to the chip. However, the NC sensors did not appear to have a different level of binding than the background binding level, indicating little or no cross-linking from the sandwich assay. Although this sensitivity is less than traditional amplification-based technologies, this device was not dependent on the rigorous heating/cooling cycles of amplification technology.
Cartridge Variability
However, these concentrations were outside the quantifiable range of the ELISA kit and therefore not as reliable. Next, we try to place a constant number of beads on the same sensor of the eight patterns. The probe is visually aligned with the surface of the sensor for each pattern and measured (Fig. 2.15).
The standard deviation of the probe measurements is significantly higher than that of blank cartridge measurements.
Limitations of dB/dz for Label-free Detection of Nucleic Acids
Although this detection method could be extremely useful due to the elimination of the need for labeling, this detection scheme is difficult to realize in frequency shift-based oscillators. The detection scheme is based on the ability to distinguish changes in the proximity of the magnetic label relative to the sensor surface. Despite this limitation, the sensing volume can potentially be used to increase the dynamic range and sensitivity of the frequency-shift biosensor by using a composite structure above the sensor surface (such as a porous membrane or a cellulose chamber) to enhance the binding of magnetic material. labels above the sensor.
Future designs could utilize the full sensor volume and increase the dynamic range of the sensor.
Additional Label-free Detection Schemes
In a variation on an earlier label-free detection scheme, the lateral sensitivity of a sensor is exploited to lead to signal change (Fig.
Variations of Magnetic Freezing
Assuming proper phase shift, the multiplication of the sinusoid and signal should correlate with beads across the sensor. Thus, the original signal is multiplied by many sinusoids with varying phase shifts, and maximum convolution power is obtained.
Absolute Oscillation Frequency of Biosensor Array
Conclusion
To use the sensor platform, a user plugs in the pre-made cartridge specific to the cartridge reader. This platform leverages research and development in magnetic bead manipulation, allowing the technology to be compatible with magnetic bead-based sample preparation. Because our reader is built with standard electronics that can be integrated into IC chips, the entire measurement system can only consist of the disposable cartridge with a small battery.
These milestones enhance the POC viability of the magnetic frequency shift biosensor and future silicon IC biosensors.
DPMF for Trafficking of Activated Macrophages for Brain Tumor Therapy
- Introduction
- Magnetic Trafficking Calculations
- Review of Magnetics
- Field Calculations from a Dynamically Controlled Grid
- Magnetic Force on a Magnetic Particle
- Magnetic Force on a Paramagnetic Particle
- Inter-particle Electrostatic Repulsion
- Fluid Dynamics of Nanoparticles and Leukocytes
- Novel Magnetic Trafficking, Proliferation, and Imaging Box
- SPION-CpG Construct
- SPION-CpG Synthesis
- Microglia SPION-CpG Uptake and Viability
- Large Scale Uptake and Distribution Imaging
- Microglia SPION-CpG Exocytosis
- Demonstration of Magnetic Response using Permanent Magnet
- Cell Viability and Morphology After Magnetic Trafficking
- Magnetic-field Induced Qualitative Cell movement in vitro by bright-
- Conclusion
Next, inductively coupled plasma mass spectrometry (ICP-MS) was used to quantify the cellular uptake of SPION-CpG by N9 microglia. Untreated control cells showed no uptake (Fig. 2.47), whereas cells treated with SPION-CpG conjugates showed profound uptake of the SPIONs (Fig. 2.47). TEM imaging was also performed to collect information on the intracellular distribution of the SPION-CpG conjugates.
N9 cells were plated on the coverslip of the cell box and then loaded with SPION-CpG.
Transistor Scaling and Variation
Self-Healing Circuits
A fundamentally different design methodology is required to ensure robust IC performance as transistor scalability and variability increase.
A Self-Healing mm-Wave Power-Mixer in 32 nm CMOS
Introduction
- Digital Backbone
- Symbol Healing
- RF Power Sensor
- DC Current Sensor
- Phase Rotator
- Phase Detection
An on-chip healing microprocessor acts as the "brain" of the self-healing circuit and automatically "heals" the chip's performance. The output of the phase shifter is directly connected to the input of the driver phases. The phase detector output versus relative phase with LO is shown in Fig. 3.25).
The output DC voltage corresponds to the phase shift of the output of the power mixer with the LO.
Variation Resistant Phase Detection Based on Min/Max Algorithm
The phase shift is a relative offset between the input of the phase detector and the LO signal. This figure shows how process variation can potentially degrade the functionality of the phase detector.
Conclusion
A Self-Healing High Power Amplifier in 32nm CMOS
- Introduction
- Self-Healing Infrastructure
- Motivation
- Surface Cleaning
- Surface Reaction
The min/max algorithm uses only the minimum and maximum output voltages of the phase detector to predict the outputs at other phase shifts. The self-healing infrastructure consists of 5 main blocks: scan chain, controller, SARADC handshake module, SARADC, an output DAC (Figure 3.33). The infrastructure interfaces with bias actuators and power sensors to adjust for a given output power/amplitude setting.
The infrastructure is designed through custom-synthesized digital logic from custom-designed standard cells (Fig. 3.35).
DNA Conjugation: Immobilization of Thiol-functionalized DNA to Amino-coated
- Motivation
- Surface Reaction
- DNA Immobilization
- Hybridization
Prepare a solution containing 95% ethanol-5% water solution (v/v) and adjust the pH to 4.5-5.5 with glacial acetic acid. Place the slides on the rack in an empty pipette tip box, fill the bottom of the box with 40mL 3x SSC and incubate for 6 h at room temperature. Immerse the slides in a solution of 50 mM 2-mercaptoethanol in 50 mM sodium phosphate buffer (pH=6.6) for 30 min to block unreacted surface-bound maleimide groups.
Immediately place the slides in 4x SSC with 0.1 wt% SDS solution, heat to 50◦C for 30 minutes to remove non-specifically bound DNA probes.
DNA Conjugation: Immobilization of Amine-functionalized DNA to Aldehyde-Coated
Motivation
DNA Immobilization
Hybridization
Detailed DNA Functionalization of IC
Materials
Protocol
Experimental Details of SPION-CpG Construct Analysis
- SPION-CpG Synthesis
- Mass Spectroscopy
- NFkB Activity Assay
- Physical Characterizations
- Inductively Coupled Plasma Mass Spectroscopy (ICP-MS) for Uptake and
- Dark-field microscopy imaging
- Electron microscopy (SEM and TEM)
- Magnetic-field Induced Qualitative Cell movement in vitro by Bright-field Mi-
- Live Cell Imaging
For each viability assay, cells were washed with PBS, treated with SPION-CpG conjugates, and incubated for 12 h. After incubation with dye, cells were treated with fixative (either glutaraldehyde for post-motion SEM samples or paraformaldehyde) and imaged. After loading, cells were washed twice with PBS and 150 µL of fresh medium was added to each well.
Cells were then loaded at 0.1 mg mL-1 SPION-CpG for 2 h, following the protocol for the previous cell motility experiment.
Optional off chip self-healing mode
Block level diagram of self-healing infrastructure
Measured results for DAC control of actuators on the power mixer (a) tail transistors
A desired constellation requires accurate phase and amplitude. A differential LO signal
RF coupler design for monitoring transmitter ouput and reflected power
Simulated coupling and isolation of RF coupler
S paramters of the couplers show little impact on output traces
RF coupler layout
RF power sensor test setup. A signal generator provides power at various levels to a
The RF power sensor’s input consists of a transistor biased at cut-off. The input
The output of two measured chips is shown. The output voltage is correlated with the
Schematic of the DC sensor
Test setup for measuring DC sensor response
Measurement results of DC sensor. The DC sensor was tested under nominal biasing
Phase rotator to aid in symbol healing. I-Q signals are generated on chip using a
Monte Carlo simulation of phase detector output over process corners. This figure
Comparison between predicted phase response using the min/max algorithm and simu-
Monte-carlo simulations of phase recovery for (a) 45 ◦ and (b) 90 ◦
Schematic of PA transistor stacking. The drain-source breakdown is 1.4V
Drain voltage scaling and drain current as a funciton of stacking
Layout of high power PA using transistor stacking
Block diagram of self-healing infrastructure
Output power and efficiency optimization algorithm
An example of a custom designed standard cell. This particular cell is a 2-input AND
Demonstration of custom synthesized digital infrastructure from custom designed dig-
