What's New 
Confocal Workshop at Dartmouth Medical School
Confocal image (Zeiss LSM510-Meta; 630X) of the mid-body section of four adult C. elegans hermaphrodites of strain CF702 muls32[mec-7::GFP;lin-15(+)]II him-5(e1490)V (a gift of Cynthia Kenyon, UCSF).
The mec-7 directed expression of GFP in the touch neuron cell bodies and their axonal projections along the anterior-posterior axis can be clearly visualized (emerald green). Gut autofluorescence, identified using the META spectral unmixing function, was distinguished unequivocally (red-gold)
Morning Session (Lectures)
- Introducing concepts of general importance in microscopy
- Introduction to confocal microscopy, sample preparation, point spread function, and appropriate sampling based on the Nyquist Theorem
- Meta detector PMY recording and characterization of spectral signatures
- Autofluorescence
- Linear unmixing of image channels
- The LSM510 acquistion software
- Co-localization analysis
Afternoon Session (Hands on Training)
- Emphasis on correct control of system parameters for COBRE project-specific components.
- Correct use of the Meta pmt detector and lambda scan acquisition will be stressed for autofluorescence imaging.
- Correct sampling frequency and control of signal cross-talk will be stressed for co-localization studies.
Specific Topics related to individual projects to be covered:
Charles Warren
All in C. elegans:
- Gut autofluorescence correction
- GFP reported axonal path tracing (neuronal processes can be long and follow intricate trajectories in a high volume sample).
- Counting the number of nuclei in a syncytial cell.
Brendan Keegan
- Tagged proteins to monitor intracellular processing, subcellular localization/ co-localization in (live) cultured cells
- Tagged mAbs to detect subcellular localization/ co-localization of proteins in cultured cells
Deena Small
- Autofluorescence correction for fat tissue
- Colocalization/ cellular localization using two different tags to see targets (i.e. V5 and B-catenin with CY3, FITC etc)
- Cytoskeletal organization: benefits of using the meta versus a regular fluorescence scope
Sophia Ouhilal
- Mouse ovarian tissues exposed to hyperglycemia (STZ) and controls and how they differ: Fluorescence at the level of granulosa and theca cells of the ovaries for glucose transporters.