Guo Research Group

We and our collaborators have also exploited the exciting nano-bio fields. We have developed a flexible technique for selectively patterning bioactive proteins using nanoimprint, surface passivation and chemical modification, and exploiting the specificity of the biotin/streptavidin linkage. This technique achieves high throughput reproducible patterns of biologically active proteins with nanoscale resolution and high selectivity (Nano Letters. 4, 953, 2004). We have exploited a specific type of motor protein, kinesin, and to integrate such nanoscopic molecular engines into functional micro-devices. We developed a highly efficient method for guiding the directional transport of microtubules by confining the kinesin proteins into passivated polymer nano-tracks that are integrated with microfluidic channels (EIPBN’2004, and Small 1, 409, 2005). We and our collaborators have obtained very encouraging results from a prototype molecular sorting device based on these developments (mTAS, 2003).  Our group also developed a greatly simplified technique for fabricating large arrays of nanofluidic channels with controlled dimensions, and illustrated one biological application by stretching genomic DNAs in the nanochannels (Nano Letters. 4, 69, 2004). Patterned arrays of nanofluidic channels may find use for single molecule detection, quick DNA sizing, integrated DNA or protein chips, or high-throughput drug screening. His group further improved the device fabrication by introducing hydrophilic HSQ material that can be imprinted at room temperature (mTAS, 2005). Our group demonstrated biomimetic polymer nanostructures that can function like the gecko palms, which can exert strong but reversible adhesions to dry surfaces (EIPBN’2006). Recently we demonstrated new approaches of ion current rectification in nanofluidic diode structures (Nano Lett. 2007). We also applied nanoimprint to the fabrication of monodispersed, position controlled and well-aligned metallic nanoparticle arrays to exploit localized surface Plasmon resonance as a label-free biosensors with very high polarization sensitivity (Adv. Mater. 2008).

 

The versatile, highly specific, and biologically friendly technique for generating ultra-high resolution protein patterns will allow the diverse activities of proteins to be integrated into microfabricated devices and sensors. It also provides a unique solution to create “grey scale” protein patterns with varied densities, which will aid the study of cell adhesion and motility and enzyme-substrate interactions. The successful demonstration of molecular sorting devices powered by motor proteins will spur further investigations and new device concepts to harvest the full potential of these highly sophisticated nanostructures provided by nature with unprecedented small scale and high efficiency. The simple technique for constructing nanochannels will provide a useful fabrication tool to perform genomic, proteomic, and chemical analysis in the nanoscale. Field-controlled nanofluidic devices may open many new applications in interfacing with cells to monitor ion fluxes. Localized surface Plasmon resonance can lead to general biosensor platforms with high sensitivity and ability to integrate with other electronics or photonics.

 

Nanofluidic Channels:

 

(a) Schematic showing the nanofluidic channel fabrication by using a template mould to imprint into a thin polymer layer to leave un-filled and self-enclosed channels. (b) SEM micrograph of imprinted nanofluidic channels with cross sections of 75 by 120 nm. (c) Fluorescent images showing the stretching of 103 kb long T5 phage DNA in the nanochannels that reaches about 95% (scale bar 50 µm).

 

 

Nanofluidic Diode

 

 

 

 

 

 

 

 

 

 

 

nanoch

 

(a) Rectifying effect due to the disparate ion distribution along a homogeneous nanochannel under different polarities of applied potential (b) Optical microscope image of a bipolar nanochannel and measured I-V characteristics of a bipolar nanochannel with bulk ion concentration varies from 10^-4 to 1M.

 

Nanoscale Protein Patterning:

Process flow diagram of substrate patterning and protein adsorption.  Spin-coated PMMA polymer is patterned by nanoimprint. Exposed SiO₂ regions are etched and a passivating (CF_x)_n polymer (x = 1 or 2, n = number of monomer subunits) is deposited during a CHF₃ RIE procedure. Residual PMMA is stripped away with acetone, exposing the underlying SiO₂ in the “patterned regions.”  An aminosilane monolayer is covalently attached to the exposed “patterned regions.  Biotin-succinimidyl ester is then covalently linked to the primary amine of the aminosilane layer, and streptavidin is bound to the biotin layer.  Finally, the biotinylated target protein is bound to the streptavidin layer.

 

 

   

(a)                          (b)                             (c)                                 (d)

Epi-fluorescence image of rhodamine-labeled streptavidin bound to uniform (a) microscale lines and (b) dots of biotinylated BSA protein on oxidized Si substrates. Fluorescent intensity signal in the passivated regions is at or below the noise level of the imaging system, indicating the fluorophore concentration in these areas is less than 0.1% of that observed in the patterned regions. Patterned streptavidin density is estimated ~120,000 molecules/µm². (c) Epi-fluorescence image demonstrating the retained biological activity of patterned biomolecules. Biotinylated polyclonal anti-catalase antibody was patterned in 2µm dots, and the antibody’s fluorescent antigen, rhodamine-labeled catalase, selectively binds to the antibody-patterned regions. The surface density of the bound catalase is approximately 31,000 molecules/µm². The scale bars are 10 mm in all three images. (d) Fluorescent micrograph of biotinylated bovine serum albumin (BSA) bound to the 75nm wide-lines pattern (SEM of the nanoline pattern is shown in the insert).

It is known that when the size of nanochannel is comparable to or smaller than the Debye length, the concentration of the counter-ions in nanochannels will be enhanced while that of co-ions diminished due to the electrostatic interaction between ions and charged channel walls. In such regime, the electric-double layers (EDL) overlap and the ion conductance through the nanochannel is determined by the surface charge instead of the bulk ion concentration in reservoirs. Our current research area include field-controlled nanofluidic devices for active control of ion polarities and concentrations. A heterogeneous nanochannel based on the opposite surface charges distributing asymmetrically along nanochannel can rectify the ion current. Recently we have demonstrated rectified ionic current through bipolar nanochannels which have opposite polarity of surface charges along 20 nm-thick, 60mm-long nanochannels. It was found that the rectification of ion current is due to an asymmetric distribution of ions with respect to the junction. Theoretical calculations agreed qualitatively with the experiments.

Bipolar Fluidic Diode

(b)

(c)