Evolve 128 EMCCD Camera from Photometrics

January 23, 2013

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University of Oxford
Physics
Doctoral Student
  • Review Image
  • Review Image
Multi-parametric imaging of voltage and calcium in a ventricular tissue-slice (350 µm-thick; adult rat heart). A) Voltage and calcium signals (normalized) are shown from the white-squared region of the tissue-slice during point electrical stimulation (at the site of the black circle). B) Normalized fluorescence intensity maps (color-bar shown) of voltage and calcium at four progressive time points, showing the progression of the activation wave. Scale-bar = 5mm.

Company:

Photometrics

Product Name:

Evolve 128

Catalog Number:

Evolve 128

ImageMuch of our research involves the development of cost-effective and simple cardiac electrophysiological imaging techniques. This field has advanced tremendously in the past three decades with developments in fluorescent dyes, optical filters, illumination sources, electronics and cameras. Here, we briefly describe a multi-parametric imaging system and its application to thin ventricular tissue slices, a low-light 2D preparation. Traditional approaches use two camera systems, which may be too costly and technically challenging for most cardiac electrophysiology research groups. We adapted the so-called ‘multi-color’ imaging technique, which relies on a single high-speed and high-performance detector, fast-switching light sources and a multi-band fluorescence emission filter. We utilized Photometrics’ Evolve™-128 EMCCD camera, modern high-power LEDs and a custom-fabricated multi-band optical filter to image both voltage and calcium fluorescence signal progression in time and space.

Experimental Design and Results Summary

Application

‘Multi-color’ imaging of thin ventricular tissue slices. The three key parameters measured were transmembrane voltage, intracellular calcium concentration and excitation wave propagation.

Starting Material

Clinically relevant changes in heart rhythm are multi-cellular phenomena. 2D tissue preparations provide a balance between simplification and representative complexity of these changes, such as life-threatening arrhythmias. In the sample results presented here, thin tissue slices (350 µm thick) from the left-ventricle of an adult rat heart were cut using a precision vibratome and submerged in physiologic solution at 37°C.

Protocol Overview

The whole-heart was loaded with voltage and calcium fluorescent dyes before the tissue-cutting procedure (in this case, di-4-ANBDQPQ and rhod-2AM, respectively). This allowed us to electrically stimulate and image voltage, calcium and excitation wave propagation shortly after cutting.

Tips

Since our system uses a single Evolve-128 camera, there were no moving parts and alignment was straightforward.

Results Summary

The figure below shows sample results from an adult rat left-ventricular tissue slice. Part A shows normalized voltage and calcium fluorescence signals, which are proportional to cell voltage and calcium transients, from one point on the slice during point electrical stimulation. Part B shows normalized fluorescence intensity maps of these parameters over time.

Features Summary

The signals in the figure below are unfiltered, highlighting the low noise level of the Evolve-128 camera. Although at full spatial-resolution the speed of the camera is high enough to capture the relevant features of the voltage and calcium transients, spatial resolution can be easily reduced via software to achieve ultra-high speeds for applications requiring more detailed interrogation of signal dynamics.

Additional Notes

None.

Image Gallery

  • Review Image
  • Review Image
Multi-parametric imaging of voltage and calcium in a ventricular tissue-slice (350 µm-thick; adult rat heart). A) Voltage and calcium signals (normalized) are shown from the white-squared region of the tissue-slice during point electrical stimulation (at the site of the black circle). B) Normalized fluorescence intensity maps (color-bar shown) of voltage and calcium at four progressive time points, showing the progression of the activation wave. Scale-bar = 5mm.

Summary

The Good

High-speed, low-noise and low-light level imaging of fluorescence signals.

The Bad

The camera system is costly.

The Bottom Line

The camera system is very well designed and versatile. With the appropriate optics, the system can be applied to microscopic single-cells and macroscopic whole-organs. Overall, the multi-parametric imaging system described here is relatively inexpensive because only a single Evolve-128 camera is used.

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