THE USE OF EXOGENOUS FLUORESCENT PROBES FOR TEMPERATURE MEASUREMENTS IN SINGLE LIVING CELLS

Photochemistry and Photobiology - Tập 62 Số 3 - Trang 416-425 - 1995
Curtis F. Chapman1, Y. Liu1,2, G. J. Sonek1,2, Bruce J. Tromberg1,3
1Beckman Laser Institute and Medical Clinic,
2Department of electrical and computer engineering
3Department of Physiology and Biophysics, University of California, Irvine, CA 92717, USA

Tóm tắt

Abstract—The fluorescent membrane probes 7‐nitrobenz‐2‐oxa‐1,3‐diazo1‐4‐y1 (NBD) and 6‐dodeca‐noy1‐2‐dimethylamino‐naphthalene (laurdan) have been studied for use as optical thermometers in living cells. The thermal sensitivity of NBD is primarily a consequence of rapid, heat‐induced electronic changes, which increase the observed fluorescence decay rate. As a result, fluorescence intensity and lifetime variations of membrane‐bound NBD‐conjugated phospholipids and fatty acids can be directly correlated with cellular temperature. In contrast, laurdan fluorescence undergoes a dramatic temperature‐dependent Stokes shift as the membrane undergoes a gel‐to‐liquid‐crystalline phase transition. This facilitates the use of fluorescence spectra to record the indirect effect of microenvironmental changes, which occur during bilayer heating. Microscope and suspension measurements of cells and phospholipid vesicles are compared for both probes using steady‐state and fluorescence lifetime (suspension only) data. Our results show that NBD fluorescence lifetime recordings can provide reasonable temperature resolution (approximately 2°C) over a broad temperature range. Laurdan's microenvironmental sensitivity permits better temperature resolution (0.1‐1°C) at the expense of a more limited dynamic range that is determined solely by bilayer properties. The temperature sensitivity of NBD is based on rapid intramolecular rotations and vibrations, while laurdan relies on a slower, multistep mechanism involving bilayer rearrangement, water penetration and intermolecular processes. Because of these differences in time scale, NBD appears to be more suitable for monitoring ultrafast phenomena, such as the impact of short‐pulse microirradiation on single cells.

Từ khóa


Tài liệu tham khảo

10.1117/12.137357

10.1364/AO.32.002439

10.1016/0009-3084(82)90053-6

10.1016/0005-2736(68)90132-6

10.1016/0009-3084(89)90066-2

10.1016/0165-022X(90)90097-V

10.1016/0165-022X(79)90043-5

10.1021/bi00544a007

10.1126/science.6849129

10.1016/0011-2240(89)90035-7

10.1111/j.1432-1033.1990.tb19114.x

10.1021/bi00075a023

10.1016/0005-2736(75)90119-4

10.1016/0005-2736(77)90078-5

10.1021/bi00623a014

10.1016/0304-4157(79)90001-7

10.1016/0009-3084(80)90028-6

10.1007/978-3-642-81537-9_8

10.1021/bi00378a027

10.1021/bi00417a005

10.1366/0003702904087325

10.1021/ac00051a018

10.1016/0304-4157(78)90010-2

10.1016/0005-2736(89)90183-1

10.1016/0009-3084(89)90049-2

10.1016/0005-2736(90)90406-E

10.1016/S0006-3495(90)82531-5

10.1016/0005-2736(73)90292-7

10.1016/0005-2736(74)90248-X

10.2307/3578478

10.1021/bi00375a006

10.1016/0009-3084(90)90128-E

10.1021/bi00297a024

10.1021/bi00517a023

10.1021/bi00541a004

10.1016/0009-3084(84)90031-8

Eidelman O., 1984, pH‐dependent fusion induced by vesicular stomatitus virus glycoprotein reconstituted into phospholipid vesicles, J. Biol. Chem., 259, 4622, 10.1016/S0021-9258(17)43092-4

10.1021/bi00364a020

10.1021/bi00365a014

10.1016/0006-291X(77)91204-9

10.1021/bi00350a001

10.1021/bi00362a023

10.1021/bi00382a008

10.1021/bi00411a009

10.1016/0005-2736(89)90276-9

10.1016/S0006-3495(92)81616-8

10.1021/bi00513a012

10.1021/bi00537a003

Nichols J. W., 1983, Resonance energy transfer assay of protein‐mediated lipid transfer between vesicles, J. Biol. Chem., 258, 5368, 10.1016/S0021-9258(20)81898-5

10.1016/0005-2736(84)90238-4

10.1021/bi00356a044

10.1021/bi00364a021

Struck D. K., 1980, Insertion of fluorescent phospholipids into the plasma membrane of a mammalian cell, J. Biol. Chem., 255, 5404, 10.1016/S0021-9258(19)70801-1

Sleight R. G., 1985, Transbilayer movement of a fluorescent phosphatidylethanolamine analogue across the plasma membrane of cultured mammalian cells, J. Biol. Chem., 260, 1146, 10.1016/S0021-9258(20)71220-2

10.1016/S0091-679X(08)60188-0

10.1083/jcb.91.3.872

Pagano R. E., 1983, Intracellular translocation and metabolism of a fluorescent phosphatide acid analogue in cultured fibroblasts, J. Biol. Chem., 258, 2034, 10.1016/S0021-9258(18)33093-X

10.1083/jcb.99.2.742

10.1083/jcb.100.1.27

10.1083/jcb.105.4.1623

Kobayashi T., 1989, Lipid transport during mitosis. Alternate pathways for delivery of newly synthesized lipids to the cell surface, J. Biol. Chem., 264, 5966

10.1016/0005-2736(86)90517-1

10.1016/0005-2736(88)90118-6

10.1111/j.1751-1097.1991.tb02028.x

10.1016/1010-6030(93)80008-W

10.1016/1010-6030(93)85048-D

10.1021/j100062a014

Parasassi T., 1986, Time‐resolved fluorescence emission spectra of laurdan in phospholipid vesicles by multifrequency phase and modulation fluorometry, Cell. Mol. Biol., 32, 103

10.1016/S0006-3495(90)82637-0

10.1021/ac00223a015

10.1016/S0006-3495(91)82041-0

10.1016/0014-4827(92)90096-Q

10.1016/0005-2736(93)90209-I

10.1016/S0006-3495(94)80763-5

10.1063/1.112981

10.1021/bi00581a025

10.1016/0005-2736(93)90399-K

Freshney R. I., 1987, Culture of Animal Cells

10.1016/B978-0-12-427150-0.50077-2

Rose G. G., 1954, A separable and multipurpose tissue culture chamber, Tex. Rep. Biol. Med., 12, 1074

Thông tin
Thông tin xuất bản

Photochemistry and Photobiology

Tập 62 Số 3

416-425

Thông tin tác giả