Nanostructures

Nanostructures are synthesized using MEMS technology. Some examples are:

• Nanowires [1]
• Nanorods [2]
• Nanoparticles [4]
• Nanobelts [5]
• Nanosheets [6]
• Nanotubes [7]
• Porous materials [8]

Each has its own advantages and disadvantages, the main idea of nanostructures is to increase the surface area ratio

Surface area ratio increase when the surface has irregularities at the surface. In the schematic from above, the addition of irregularities onto the surface tends to create more free places for the recognition bio molecules to attach.

Many different nanostructures can be deposited on the sensor working area. Nanowires or nanotubes can be aligned, nanorods and nano particles usually enhance the sensitivity by attaching themselves to biomolecules and thus enhancing electrical conductivity, nanobelts as well as Nanosheets can provide several new layers for adhesion, and finally porous materials (like porous Silicon) can also act like a filter by only letting the correct size particles through it. The decision on how to use a certain nanostructure depends entirely on creativity and experience.

[1] Fabrication of nanowires of multicomponent oxides: Review of recent advances, Shankar K. S, Raychaudhuri A. K, Materials science and engineering C, 25, 738-751(2005)
[2] Gold nanorods: Synthesis, characterization and applications, Pérez-Juste J, Pastoriza-Santos I, Liz-Marzán L. M, Mulvaney P, Coordination Chemistry reviews, 249, 1870-1901(2005)
[3] Manufactured nanoparticles: An overview of their chemistry, interactions and potential environmental implications, Ju-Nam Y, Lead J. R, Science of the total environment, 400, 396-414(2008)
[4] Nanobelts of Semiconducting Oxides, Pan Z. W, Dai Z. R, Wang Z. L, Science, 291, 1947-1949(2001)
[5] Preparation and characterization of graphite nanosheets from ultrasonic powdering technique, Chen G, Weng W, Wu D, Wu C, Lu J, Wang P, Chen X, Carbon, 42, 753-759(2004)
[6] Nanostructuring electrode with carbon nanotubes: A review on electrochemistry and applications for sensing, Gooding J. J, Electrochimica Acta, 50, 3049-3060(2005)
[7] Properties of porous silicon nano-explosive devices, du Plessis M, Sensors and Actuators A, 135, 666-674(2007)
 
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B-Cells and antibodies

B cells are specialized immune cells that can detect virtually any kind of pathogen and produce millions of antibodies. When a B cell detects a pathogen, it will then evolve into a plasma (producing antibodies) or memory cell (recording the pathogen for future attacks)

Schematic of B cell differentiation (usually a little more complicated than this), B cells (naive)  creates antibodies at their surface, they become activated when a specific antigen (it can be a virus, a bacteria, or another unidentified particle, here exemplified as the green star), gets recognized by the antibody. With the help of other cells (T helper cells), they can evolve into plasma B cells (producing millions of antibodies, with a shorter life span) or a memory cell (a cell that remembers the type and specificities of the antigen, but with a longer life span)

How science uses antibodies?
Because of their ability to detect only one specific biomolecule, antibodies are often viewed as the best way to interact with biosensors.

A strategy to mass produce them is the Hybridoma technology (developed in 1975 by Milstein, and Köhler). Briefly, a plasma B cell is fused together with a cancer cell, in this way, the plasma B cell does not die and will keep on producing millions of specific antibodies. [1]

A simplified overview of an antibody(IgG), showing the principal structures. In human bodies, they can conjugate with antigens and facilitate macrophage eating. It also can initiate lysis in cooperation with the complement attack complex

Antibodies work well with ELISA (Enzyme Linked Immuno Sorbent Assay) where the antibodies detect a specific antigen and they are linked next with a fluoro-enzyme(an enzyme capable of emitting light), a deep analysis on a microscope will yield the response, if everything goes well, the assay will be as bright as a field of fireflies.

Antibodies have been also used in cantilevers or membranes (MEMS) to detect by weight the presence of a given antigen. [2] The detection strategy in this case, can be optical, piezoelectric, or even by capacitance.

Cantilever with antibodies, based on the size and weight, the cantilever can deflect downside or even upside as seen on [3] (with SCFV instead of antibodies)

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[1] Human antibodies by design, Vaughan, T. J, Osbourn, J. K, Tempest, P. R, Nature biotechnology, 16, 535-539 (1998)
[2] Cantilever transducers as a platform for chemical and biological sensors, Lavrik. N. V, Sepaniak, M. J, Datskos P. G, Review of scientific instruments, 75, 2220-2253 (2004)
[3] A label-free immunosensors array using single-chain antibody fragments, Backmann N, Zahnd C, Huber F, Bietsch A, Plückthun A, Lang H. P, Güntherodt H. J, Hegner M, Gerber C, PNAS, 102, 14587-14592 (2005)

Amino acids and proteins

Amino acids are the basic building blocks of all proteins. The human body requires at least 20 amino acids to produce different proteins, but the human body can only produce 10 of them, the other are found in our daily food income (healthy diet only).
The protein production starts at the nucleus of each human cell. And they serve the cell by executing a vast physical or chemical tasks. All of these tasks are determined by our own DNA and the way that a protein envelopes itself.
For a more detailed information please follow the links:
Animation of proteins and amino acids
Basics of amino acids

Basics on biosensing

Introduction

What exactly is a Biosensor?
Sensing biomolecules is the main reason of  the existance of biosensors, they include a biorecognition element, a transducer, and electronics circuits to generate a readable output.

Schematic of a Biosensor, a simplified one here. From top to bottom: The analytes (the biomolecule that we are interested in), are plenty and differ  in shape and size,  from circles to crazy star shaped things. Take a closer look and you will find only one element that fits correctly (also has the same color), to the biorecognition element. The transducer here is simplified as a the little yellow box, and a line to the read out system is coming from it. This in real life is often more complicated, and the readout system can be sometimes without the use of any electronic circuitry.

Biomolecules can be:
Biomolecules are large biological constructions. These large constructions are made up of amino acids, their function range from (some examples):

• Being part of the outer layer structure of cells, some cells can have a lipid layer structure that will impede water from entering the cell, in viruses for example, their outer layer keeps their genetic information protected, and help them infect other cells or bacteria.

• They can transport other biomolecules, some proteins(others do different things), can take peptides and transport them inside the cell.

• Some can even detect viruses or bacteria, antibodies are glycoproteins made up by B-cells, capable of detecting harmful organisms, there can be an infinite number of different antibodies each one detecting a different antigen.

• They can also gather biological information, DNA keeps all the genetic information stored, and it only needs some billions aminoacids.

The biorecognition element is most of the time, a biomolecule (an antibody, a SCFV,  an Aptamer or an enzyme), that will only detect the analyte, for example: The biorecognition element for detecting PSA is anti-PSA, another example is: Glucose oxidase (an enzyme), that will only recognize Glucose.
Learn more about aminoacids and the different biomolecules that they can form

What can Nanotechnology do for our biosensors?
Nanotechnology can improve most biosensors by increasing the contact area (the surface-area to volume ratio), depending of the material used, some nanostructures can also enhance their electrocatalytical effects.
Learn more about Nanostructures.

How can Nanotechnology is integrated into biosensors?
Chemically modifying the surface of the substrates can introduce the correct linkers for the nanostructures to get immobilized. This depend on the material, and crystal orientation.
Usually all biomolecules are either amino or carboxyl group terminated

What are the future trends in Biosensors?
Right now, most biosensors detect their analytes by detecting them outside the human body. This can pose many problems as the body also recognizes the sensor as foreign and the immune system attacks it, also, some of the materials used are often toxic to different organisms.
Different trends suggest the need of an implantable biosensor, with the following enhancements:

• Self sufficient (wise energy harvesting)
• Able to communicate with the outside (for monitoring purposes)
• Bio-Compatible (non toxic and immuno friendly)
• Stable, cheap, easy to produce (environmentally amicable) 

What is the current work in USF?
In the University of South Florida, many different biosensors have been created.
A common example, integrating Nanotechnology in biosensors in the USF.
 
Special thanks for the fulfillment of this work goes to the Bio-MEMS group in USF. Kind acknowledgments goes to Dr. Sunil Kumar Arya for the chemical teaching, Eric Huey and Dr. Subramaniam Krishnan for growing the Silicon Oxide Nanowires.  

Something to read about:
[1] Biosensor Recognition Elements, Chambers J. P, Arulanandam B. P, Matta, L. L, Weis A, Valdes J. J, Current issues in Molecular Biology, 10, 1-12(2006)
[2] Aptamer-based biosensors Song S, Wang L, Li J, Zhao J, Fan C, Trends in Analytical Chemistry, 27, 108-117(2008)
[3] Antibody phage display technology and its applications, Hoogenboom H. R, de Bruïne A. P, Hufton S. E, Hoet R. M, Arends J. W, Roovers R. C, Immunotechnology, 4, 1-20(1998)
[4] Selecting and screening recombinant antibody libraries, Hoogenboom H. R, Nature biotechnology, 23, 1105-1116(2005)
[5] Manipulating redox systems: Application to nanotechnology, Gilardi G, Fantuzzi A, Trends in Biotechnology, 19, 468-476(2001)
[6] Surface modification in microsystems and nanosystems, Prakash S, Karacor M. B, Banerjee S, Surface science reports, 64, 233-254(2009)

[5] Electrochemical Glucose Biosensors, Wang J, Chem. Rev, 108, 814-825(2008)

Biosensing in USF

Glucose measurements involve a large variety of sensors and methodologies.[1-2] They have been around since 1967 when Updike and Hicks created the first glucose sensor.
The relevance of creating Glucose sensors is due to the problematic incidence of blood-sugar levels. One such problem is diabetes. Normal Glucose in blood ranges from 85 to 135 mg/dl, 
In the University of South Florida, working with enzyme biosensors is a good way to be introduced to the marvelous world of both nanotechnology and biology.
In order to create Silicon Oxide Nanowire glucose sensors, one needs:

1. Silicon oxide Nanowires.[4]
2. Electrodes.
3. Chemical reagents.
4. Glucose Oxidase.
5. Glucose.
6. Electronic circuitry.

Process on creating the electrodes. 1.- Starts with the creation of the Silicon Oxide Nanowires at the CVD.[4] 2.- Exemplifies the separation and posterior adhesion of nanowires to the electrodes. 3.- Chemical modification so the Glucose Oxidase can be integrated to the system.

In USF we use electrochemical experiments to detect glucose in various concentrations. Silicon oxide nanowires are first grown by Chemical Vapor Deposition (CVD).[4]

Silicon oxide in solution, Scanning Electron Microscope at USF

Then, they are deposited onto gold electrodes (glass substrate)

Gold electrodes with different amount of Silicon nanowires, the higher concentration at the left and the lower concentration (no silicon nanowires) to the right.

After Silicon oxide deposition, a chemically modified Glucose Oxidase (GOx) is attached to the nanowires creating the appropriate physicochemical structure that can detect Glucose.

Silicon Oxide Nanowires after deposition over the gold electrode, Focused Ion Beam at USF

Silicon Oxide Nanowires with Glucose Oxidase attached, note that its only attaching to the nanowires, Focused Ion Beam at USF

Electrochemical techniques such as Cyclic voltammetry(results not shown here), are used to detect glucose concentrations. [3] The main idea here is that the enzyme, acting as an electrocatalytical agent, [5] will modify glucose and for each modification we will have more electrons in the system. The larger the quantity of glucose in solution, the larger the increment in current that will be detected.

For more information on related topics, please read:

[1] Glucose monitoring: state of the art and future possibilities, Wilkins E, Atanasov P, Med. Eng. Phys, 18, 273-288(1996)
[2] Home blood glucose biosensors: a commercial perspective, Newman J. D, Turner, A. P. F, Biosensors and Bioelectronics, 20, 2435-2453(2005)
[3] Status of biomolecular recognition using electrochemical techniques, Sadik O. A, Aluoch A. O, Zhou A, Biosensors and Bioelectronics, 24, 2749-2765(2009) 
[4] Manufacturing aspects of oxide nanowires, Sekhar P. K, Bhansali S, Materials Letters, 64, 729-732(2010)
[5] Glucose oxidase from Aspergillus niger: the mechanism of action with molecular oxygen, quinones, and one-electron acceptors, Leskovac V, Trivić S, Wohlfahrt G, Kandrac J, Pericin D, IJBCB, 37, 731-750(2005)

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