Study On Intracellular Ca～(2+) Concentration With Ratio Fluorescence Image Master

Calcium as a second messenger transfers external signal ( hormone, light , temperature and other environal stimulates) to intracellular signal, and induces a series of intracellular affairs. Abundant researches indicated that the messenger function of Ca2+ is realized by adjusting and controlling the intracellular Ca2+ concentration. The produce and end of Ca2+ signal resulted inadd-and-subtract and surge of intracellular Ca . So the measurement of intracellular Ca2+ concentration is very important.The development of new fluorescence dye and imaging technique have greatly promoted the research in the field of cellular calcium signal transduction path. In the first part of this thesis, the history and development of the Ca2+ fluorescence measurement were introduced, the function of Ca2+ as intracellular second messenger in the signal transduction path and the theory of cellular calcium fluorescence measurement were presented and our experiment system was reported. Research on the ratio method of measuring intracellular Ca2+ concentration with ratio fluorescence ImageMaster was reported in the last chapters, which include the folio wings:(1) The ratio experiment system of measuring intracellular Ca concentration with ratio fluorescence ImageMaster was established;(2) The primary culture mode of suckling mouse cardiac muscle cell was established and the Ca2+ concentration in suckling mouse cardiac muscle cellwas measured. After adding NE(norepinephrine ) to cardiac muscle cell ,the relative fluorescence intensity was dramatic increased. The phenomenon agreed with physiological action feature of ME. So our experiment system was proved. The change of intracellular Ca2+ concentration can be measured after adding different medicine with our experiment system. And the mechanism of action of medicine to cell can be researched form biomedicine;(3) The Ca2+ concentration in B16 cell was measured with ratio fluorescence ImageMaster. The preparation of cell and the carry method of fluorescence indicator were confirmed; the acquisition of cell image and the ratio analysis method by software-ImageMaster were established; the acquisition of cell fluorescence image and the ratio analysis method by software-Felix were established.