Confocal Microscopy

Microscopy Core Facility

Overview

The Microscopy Core Facility (MCF) provides confocal microscopy services to support both research and clinical applications.

Confocal microscopy enables high-resolution imaging and optical sectioning of biological specimens, allowing users to analyze structures within cells and tissues.

Leica TCS SP8 Confocal Microscope

The Leica TCS SP8 confocal microscope is an inverted, laser-powered, single-point true confocal scanning system. It allows users to produce high-resolution, publication-quality images of fluorescently labeled, histologically fixed tissues and cells, as well as cultured live cells.

System Capablities and Specifications

  • The system includes the following capabilities:
    • True confocal scanning system (TCS) with switchable non-resonant and resonant scanning
    • Scanning speeds up to 84 frames per second in standard mode and up to 290 frames per second in resonant mode
    • Maximum resolution of 8192 x 8192 pixels (standard mode) and 1024 x 1024 pixels (resonant mode)
    • Optional field rotation up to 200 degrees using a König-Rotator
    • Adaptive Focus Control (AFC) to maintain focal plane stability during long-term experiments
    • Spectral detection range from 400 nm to 800 nm
    • Up to five individually regulated detection channels

Imaging Technology

  • The system uses advanced optical components to improve imaging quality:
    • Pellin-Broca prism design for high spectral transparency
    • Leica LIAchroic beam splitters for improved contrast and transmission
    • Hybrid (HyD) and photomultiplier tube (PMT) detectors for efficient signal detection
    • Brightfield transmitted light detector

    These features support high-contrast imaging while reducing photobleaching and preserving sample integrity.

Live Cell Imaging

The system includes a live cell incubation chamber that supports long-term imaging of live cell cultures.

  • The chamber maintains:
    • Proper carbon dioxide (CO2) levels
    • Stable temperature
    • Controlled humidity

    This allows for extended experimental conditions and time-lapse imaging.

Objectives and Imaging Options

  • Available microscope objectivies include:
    • 10x/0.40 (dry)
    • 20x/0.75 (dry)
    • 40x/1.30 (oil)
    • 63x/1.40 (oil)
    • 100x/1.40 (oil)

    Differential Interference Contrast (DIC) is available with the 20x, 40x, 63x and 100x objectives to enhance image contrast.

Laser Configuration

  • The system includes four solid-state lasers:
    • 405 nm
    • 488 nm
    • 552 nm
    • 638 nm

Workstation and Software

  • The microscope is supported by a high-performance workstation with:
    • Windows 10 Professional (64-bit) operating system
    • Intel Xeon processor
    • NVIDIA Quadro GPU
    • 6 TB storage capacity
    • 30-inch high-resolution monitor

    Leica LAS X software is used for image acquisition and analysis.

    Additional standalone computers with Leica LAS X software are available for data viewing and analysis.

Reservations

To reserve the confocal microscope, please email Rolf Fritz at fritzr@etsu.edu.

Additional Information

  • Usage Fees:

    ETSU and Educational Users:

    • $25 per hour (assisted use)
    • $25 per hour (unassisted use)

    Commercial Users:

    • $120 per hour (assisted use)
    • $80 per hour (unassisted use)
  • Policies and Procedures

    All users must follow these policies:

    • Users must be trained before unassisted use
    • Operating procedures are available in the confocal room
    • Users must complete a sign-up sheet and provide a valid billing account number
    • Users must bring their own storage device for data
    • Data is not guaranteed to be stored long-term on facility computers
    • No food or drink is allowed in the room
  • Data and Publication Acknowledgment
    Users are expected to acknowledge the Microscopy Core Facility in publications and professional manuscripts when data obtained from the facility is used.