Calcium Imaging Microscope

This is an Olympus BX61 WI fixed-stage motorized upright microscope. It has a built-in motorized Z-drive (0.01 µm resolution) with motorized brightfield and fluorescence illuminator. Fluorescence is capable of producing light output from 340 nm to a cutoff of 700 nm. Six-position motorized filter turret as well as manual fluorescence attachments are available for fluorescence scan. An arc lamp illumination source for maximum excitation wavelength flexibility. 4X dry, 40X dry and 60X water objectives are currently available for observation. Slide bookTM 4.2 is employed to record and analyze the images.

Applications

Intravital experiments

This upright microscope has a water immersion (WI) line of objectives with long distance, large approach angles for manipulator access, which is ideal for intravital experiments.

Calcium imaging

The microscope is equipped with 340 and 380 nm excitation filter wheel, which allows calcium ratio imaging after FURA-2 loading at 5 frames per second (fps) at full resolution (see sample image). Binning allows faster frame rates. In addition, filters for GFP, CFP, YFP, PE, Cy5, DAPI and Fura-2 are installed in this system. This is achieved by the Chrom A 88000 filter set, which can show DAPI, Violet, FITC/GFP, Texas Red and Cy5. The other set of filters is Chrom A 71000V2 which consist of 340/380 excitation and 515 (GFP) emission. The 40X dry and 60X water objective are available for acquisition the images.

Location: First floor BSLII suite, Room 4.

Contact information: Hong Zhang, IB-Ley lab

Training procedures contact information: Leo Fernandez

Figure 1. Microscope, approximately as installed.


Figure 2. Mouse neutrophil in a flow chamber, flow from right to left. Intracellular free calcium concentration measured by FURA-2 as ratio of 340/380 nm excitation, encoded by pseudocolor (blue is low, red is high).