About

MBIC Fluorescent Biosensors Overview

The Molecular Biosensor and Imaging Center is an association of biologists, chemists, and engineers functioning within the Mellon College of Science. Over the past decade we have developed detection technologies based primarily on fluorescence methods. Projects included a collaboration funded by NASA with a field robotics team to detect life in extreme environments, cooperative research on imaging heart excitation to follow cardiac function, development of multi-mode fluorescent MRI contrast agents, design and synthesis of quantum dot derivatives for the mesoscopic fluorescence imaging of small animals, and the development of reagents and methods in bone tissue engineering.

More recently, the MBIC has been selected as a National Technology Center to develop Fluorescent Biosensors for Networks and Pathways. The goal of this research is to create tools to obtain spatial and temporal information about the structure and function of pathways in living cells. Hundreds of regulatory proteins interact dynamically in the three-dimensional space of individual living cells to control normal and disease activity. Novel sensors of molecular interactions are needed to detect, in real time, these association and dissociation processes.

Basics of FAPs and Fluoromodule Technology

Six years ago we received major NIH center funding to develop a new concept for biosensors of protein interactions. We have identified dyes, fluorogens, whose fluorescence properties are dependent on their environment. We have also shown that single chain antibody (scFv) proteins can be selected which bind with fluorogens to produce fluorescent biosensors. These fluorogens are engineered dyes, the engineered antibody fragments are "fluorescence activating peptides", or FAPs; the combined fluorogen-FAP biosensors we term fluoromodules.

This fluorogen-FAP technology is strikingly effective for studying processes in living cells. The dyes do not fluoresce until they are constrained by binding to their FAP partners and therefore alone the dyes give no background fluorescence. The true power of this concept, however, arises from the protein character of the FAP scaffolds. Cells can be engineered to express these FAPs either as independent peptides or fused with another protein of interest. The presence of fluorogen will only generate fluorescence signal in those cells actively expressing the FAP peptide.

Examples of validated biosensors we are actively studying:

  • Biosensors for selectively quantifying densities of labeled membrane proteins and tracking their endosomal pathways.
  • Biosensors for measurement of live cell intracellular and extracellular protease activities.
  • Biosensors for quantifying the pH environment of labeled proteins as they move through cells.
  • In addition we are developing additional important biosensor classes:

  • Targeted physiological probes for membrane potential, calcium, sodium, potassium, and redox potential of selected regions or compartments of living cells or targed for selected cells in a complex population.
  • Biosensors that provide instantaneous readout of protein expression.
  • A powerful biosensor family (Lightup Antibodies) in which the intracellular biosensor becomes fluorescent when it detects (transiently associates with) specific molecular structures within cells such as a phosphorylated peptide sequence vs. a non-phosphorylated sequence.
  • A biosensor to visualize dynamic protein-protein interactions in living cells.
  • MBIC


    The Molecular Biosensor and Imaging Center at Carnegie Mellon University is a collaboration of biologists, chemists, and engineers developing fluorescence technologies for biomedical applications. The Center is funded as a National Technology Center by the National Institutes of Health Common Fund's Building Blocks, Biological Pathways, and Networks program.

    CMU


    Carnegie Mellon University (CMU) is a global research university with more than 13,200 students, 100,000 alumni and 5,000 faculty and staff.


    CMU has been a birthplace of innovation since its founding in 1900. Today, we are a global leader bringing groundbreaking ideas to market and creating successful startup businesses.

    History


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    The Molecular Biosensor and Imaging Center (MBIC), at Carnegie Mellon University has existed in some form as a Center for almost 30 years and is now directed by Marcel Bruchez, PhD. The National Institutes of Health named us a National Technology Center for Networks and Pathways, a collaboration among Carnegie Mellon University, the University of Pittsburgh (The Center for Biologic Imaging and the Department of Pharmacology), The University of California at Berkley and Stanford University. The Center will focus on the development of optical biosensors and imaging informatics for the detection of molecular interactions within living cells.

    The Center, originally named The Center for Fluorescence Research was founded in 1982 by D. Lansing Taylor, PhD. In 1991, it was renamed The Center for Light Microscope Imaging and Biotechnology, joining fluorescent probe technologies and computerized fluorescence microscopy to study temporal and spatial interactions of living cells. The Center was a National Science Foundation Science and Technology Center for 11 years with innovative research, technology transfer, and outreach and education programs. It has pioneered automated, multicolor fluorescence imaging technologies that have now been adopted by Carl Zeiss, Nikon, Olympus and other major imaging microscope manufacturers. The Center is also famous for its development of the CyDye labeling technologies that are used worldwide in biomedical research and diagnostics in place of radioisotopes. The center is particularly strong because of the interdisciplinary collaboration of its scientists, engineers and medical doctors. This interdisciplinary research brings international prestige to the center. For example, in June 1996, the center was honored with the Computerworld Smithsonian Award for Science and Innovation, one of the most prestigious awards for scientific innovation. The Center has continued with funding from NIH, NSF, NASA, the Keck, Moore and Fine Foundations.

    MBIC, established in 2000 by Prof. Alan Waggoner, functions within the Mellon College of Science of Carnegie Mellon University. Laboratory facilities include organic chemistry, imaging microscopy, flow cytometry (both analytical and high-speed sorting), cell biology, biochemistry, tissue culture, and instrumentation development. Available imaging microscopy systems include a Carl Zeiss LSM 510 Meta NLO confocal microscope with Confocor 3 FCS module, a Carl Zeiss LSM 510 Meta UV DuoScan confocal microscope, a home-built Olympus FluoView 300 multi-photon confocal microscope, an Olympus spinning disk confocal microscope, a Carl Zeiss ApoTome fluorescence microscope, a Carl Zeiss fluorescence microscope with FemtoJet microinjector, a Carl Zeiss fluorescence microscope with single molecule sensitive CCD camera and two Carl Zeiss long-term live cell time lapse fluorescence microscopes.. HPLC equipment, absorption and fluorescence spectrometers are also available. Mass spectroscopy, NMR, IR and other analytical tools are available in Chemisty Department facilities in our building.

    Our Management Team


     
    • Connector.

      Marcel Bruchez

      Director

    • Connector.

      Donna Smith

      Program Coordinator

    • Connector.

      Robert Bodnar

      Business Manager

     

    Contact US


    Molecular Biosensor and Imaging Center
    Carnegie Mellon University
    4400 Fifth Avenue
    Pittsburgh, Pennsylvania 15213

    Phone: 412-268-3461
    Fax: 412-268-6571