Research Tools

Most Recent Inventions

Efficient In Vitro Assay for Antigen-Specific Tolerance

Building on their work, UW–Madison researchers have now developed a T cell-bound cytokine (T-CBC) assay for detecting and quantifying regulatory T cells specific to self-antigens or donor alloantigens. The new method comprises (a) culturing the subject’s T cells for 24 hours in the presence of one or more target antigens and (b) analyzing the cultured T cells for expression of a marker (EBi3; TGFβ/LAP) indicative of antigen-specific immune suppression.

Monomeric Fluorescent Protein-Ligand Complexes with Strong Fluorescence in the Far-Red Region

Research from the University of Wisconsin-Washington County in collaboration with the Institute for Stem Cell Biology and Regenerative Medicine in India, has resulted in the development of monomeric variants of the naturally occurring Sandercyanin Fluorescent Protein (SFP) using site-directed mutagenesis. This work has stemmed from earlier research focused on development of the tetrameric form of SFP, a biliverdin-binding lipocalin protein originally isolated from the mucus of the blue walleye fish, Sander vitreus. Monomeric variants of SFP (mSFPs) have been found to possess the same non-covalent, bili-binding characteristics of the tetramer but are one-quarter the size (~18.6kDa) and do not oligomerize. They are therefore anticipated to be more useful in a host of biotechnology applications. Like the tetrameric form, the mSFPs have a large stokes shift (375nm/675nm) and fluoresce in the far-red or near infrared region, which is advantageous for a wide range of applications including investigation of protein-protein interactions, spatial and temporal gene expression, assessing cell biology distribution and mobility, studying protein activity and protein interactions in vivo, as well as cancer research, immunology, and stem cell research and sub-cellular localization. In addition, the newly developed mSFP’s far-red fluorescence is particularly advantageous for in vivo, deep-tissue imaging.

Enhanced Endotoxin Detection: New Advantages in Liquid Crystal Assays for Gram-Negative Pathogens

UW–Madison researchers have now demonstrated enhanced endotoxin detection in the presence of masking agents in their previous liquid crystal system.

Unlike the LAL assay, the LC-based method does not suffer from LER or any loss of sensitivity due to the presence of cations (e.g., Ca2+ or Mg2+), buffers (e.g., citrate), surfactants (e.g., SDS), chelating agents (e.g., EDTA), proteins or nucleic acids (e.g., DNA or RNA). Thus, the LC-based method provides faster and cheaper detection of endotoxin when compared to existing methods, such as the LAL assay.

Adapted Rhinovirus C for Maximum Virus Yield

Building on their work, the researchers have now developed a mutated RV-C strain that induces strong cytopathic effect and replicates vigorously in the HeLa-E8 cells, yielding more than a log higher level of infectious rhinovirus particles compared to the parental clinical isolate.

DNA “Millichip” Enables Low-Cost, High Throughput Gene Expression Analysis

UW-Madison researchers have developed a DNA “millichip” designed for low-cost, high throughput gene expression analysis in whole genomes.  The millichips consist of 1,000 to 100,000 different oligonucleotides probes immobilized on small solid support arrays with relatively high density.  The probes, which range from 30 to 100 nucleotides long, occupy separate, known sites in the arrays. 

For example, a maskless array synthesizer (MAS) can be used to synthesize about 800,000 70-mer oligonucleotides on a glass microscope slide.  Then the slide is divided into 96 pieces, each containing about 30,000 of the 70-mer DNA sequences.  These small pieces can be used in any experiment that uses standard DNA chips.

Because the millichips are small, less than 10 cubic centimeters, small volumes of solutions can be used for analysis.  In addition, the small substrate size allows the arrays to be visualized using instrumentation readily available in research laboratories.

Most Recent Patents

Yeast-Based Intein Platform for Drug Production

UW–Madison researchers have engineered non-self-cleaving Mxe GyrA inteins shown to significantly improve the production of fusion proteins from Saccharomyces cerevisiae. The novel inteins were developed through directed evolution, and they enhance fusion protein display (up to 3x) and secretion levels (up to 30x) compared to the wild type intein. The new yeast-based platform provides a robust alternative to bacterial intein expression systems.

Hydrogel Arrays for Screening Cell-Substrate Interactions, Now in Multiwell Format

Building on their previous work, the researchers have now adapted their method to any commercially available, glass or polystyrene-bottom multiwell plate. In the new process, hydrogel is covalently immobilized to the bottom of each well and then selectively polymerized. In this way the spots are completely isolatable, allowing for systemic and independent control of their chemical composition and XYZ physical dimensions.

Once the hydrogel array is formed, each of the spots can be exposed to different soluble factors without risk of diffusion.

Robust Substrates Expand the Utility of Surface Plasmon Resonance Imaging for Analysis of Biomolecular Interactions

UW–Madison researchers have developed robust, SPR-compatible substrates. The key to these substrates is a rugged, chemically versatile carbon thin film overlayer placed on an SPR-active metal thin film.

Specifically, the substrates include a support surface capable of transmitting light, a metallic layer adhered to the support surface and a carbonaceous layer deposited on the metallic layer. The substrates also may include biomolecules attached to the carbonaceous, or carbon-rich, layer. These biomolecules may include oligonucleotides, DNA, RNA, proteins, amino acids, peptides or other small biomolecules that can be configured in one or more arrays.

The new substrates are more robust than conventional gold substrates, allowing assays to be performed under higher temperatures and harsher chemical conditions than currently is possible. Additionally, the carbon thin film overlayer is not susceptible to damage from UV irradiation.