Pflügers Archiv

, Volume 409, Issue 1–2, pp 47–51 | Cite as

An inexpensive dual-excitation apparatus for fluorescence microscopy

  • John Kassotis
  • Susan F. Steinberg
  • Samuel Ross
  • John P. Bilezikian
  • Richard B. Robinson
Excitable Tissues and Central Nervous Physiology

Abstract

Ion sensitive indicator molecules can be employed in conjunction with fluorescence microscopy in single cells to measure rapid changes in the intracellular concentration of several ionic species. A number of these probes (e.g. fura-2) require the capability of measuring emission intensity at two excitation wavelengths to quantitate properly intracellular ion concentration. We have developed a simple dualexcitation apparatus for use in such applications. The apparatus switches the excitation filter within 150 ms. This economical apparatus is well suited in situations where the ionic concentration of interest is changing relatively slowly. Moreover, by synchronizing the device's action with an external stimulus; rapid and reproducible ionic changes in excitable tissue also can be measured.

Key words

Fura-2 Fluorescence microscopy Dualexcitation Instrumentation Excitable tissue Ion sensitive probe 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Grynkiewicz G, Poenie M, Tsien RY (1985) A new generation of Ca2+ indicators with greatly improved fluorescence properties. J Biol Chem 260:3440–3450Google Scholar
  2. Hermsmeyer K, Robinson KB (1977) High sensitivity of cultured cardiac muscle cells to autonomic agents. Am J Physiol 233:C172-C179Google Scholar
  3. Hewett K, Legato MJ, Danilo P Jr, Robinson RB (1983) Isolated myocytes from adult canine left ventricle: Ca2+ tolerance, electrophysiology, and ultrastructure. Am J Physiol 245: H830-H839Google Scholar
  4. Paradiso AM, Tsien RY, Machen TE (1985) Na+-H+ exchange in gastric glands as measured with a cytoplasmic-trapped fluorescent pH indicator. Proc Natl Acad Sci USA 81:7436–7440Google Scholar
  5. Ryan US, Clements E, Habliston D, Ryan JW (1978) Isolation and culture of pulmonary artery endothelial cells. Tissue Cell 10:535–554Google Scholar
  6. Williams DA, Fay FS (1986) Calcium transients and resting levels in isolated smooth muscle cells as monitored with quin 2. Am J Physiol 250:C779-C791Google Scholar

Copyright information

© Springer-Verlag 1987

Authors and Affiliations

  • John Kassotis
    • 1
  • Susan F. Steinberg
    • 1
  • Samuel Ross
    • 1
  • John P. Bilezikian
    • 1
  • Richard B. Robinson
    • 1
  1. 1.Departments of Pharmacology and MedicineColumbia University, College of Physicians and SurgeonsNew YorkUSA

Personalised recommendations