Terbium chelate membrane label for time-resolved, total internal reflection fluorescence microscopy of substrate-adherent cells

被引:32
作者
Phimphivong, S [1 ]
Saavedra, SS [1 ]
机构
[1] Univ Arizona, Dept Chem, Tucson, AZ 85721 USA
关键词
D O I
10.1021/bc9701609
中图分类号
Q5 [生物化学];
学科分类号
071010 ; 081704 ;
摘要
A chelating agent conjugated to a lipid was synthesized by reacting the dianhydride form of diethylenetriaminepentaacetic acid sequentially with 4-aminosalicylate and dioleoylphosphatidylethanolamine. The product, DOPE-YAS-Tb, exhibits photophysical properties characteristic of a chelated Tb3+ ion bound to an organic triplet donor: an excitation maximum at 310 nm, narrow emission bands at 490, 545, 590, and 625 nm, and a lifetime of 1.57 ms. The suitability of DOPE-YAS-Tb as a membrane-staining agent for morphological studies of cultured cells using total internal reflection fluorescence microscopy (TIRFM) was investigated. Swiss albino mouse 3T3 cells were cultured on silica and polystyrene substrates. Time-resolved detection was employed to reject short-lived background emission (autoemission from the cells and/or the polymer substrate), which allowed the long-lived Tb3+ emission to be selectively imaged. The results show that time-resolved TIRFM of cells stained with DOPE-YAS-Tb is an effective method of quantitatively examining the cell morphology in situations where background due to autoemission from cells and/or the substrate material is problematic.
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页码:350 / 357
页数:8
相关论文
共 31 条
[1]   AUTOFLUORESCENCE OF VIABLE CULTURED MAMMALIAN-CELLS [J].
AUBIN, JE .
JOURNAL OF HISTOCHEMISTRY & CYTOCHEMISTRY, 1979, 27 (01) :36-43
[2]   CELL-SUBSTRATE CONTACTS ILLUMINATED BY TOTAL INTERNAL-REFLECTION FLUORESCENCE [J].
AXELROD, D .
JOURNAL OF CELL BIOLOGY, 1981, 89 (01) :141-145
[3]   TERBIUM CHELATE FOR USE AS A LABEL IN FLUORESCENT IMMUNOASSAYS [J].
BAILEY, MP ;
ROCKS, BF ;
RILEY, C .
ANALYST, 1984, 109 (11) :1449-1450
[4]   MICROFILAMENTS AND ACTIN-ASSOCIATED PROTEINS AT SITES OF MEMBRANE SUBSTRATE ATTACHMENT WITHIN ACETYLCHOLINE-RECEPTOR CLUSTERS [J].
BLOCH, RJ ;
VELEZ, M ;
KRIKORIAN, JG ;
AXELROD, D .
EXPERIMENTAL CELL RESEARCH, 1989, 182 (02) :583-596
[5]   QUANTITATIVE-ANALYSIS OF VARIABLE-ANGLE TOTAL INTERNAL-REFLECTION FLUORESCENCE MICROSCOPY (VA-TIRFM) OF CELL SUBSTRATE CONTACTS [J].
BURMEISTER, JS ;
TRUSKEY, GA ;
REICHERT, WM .
JOURNAL OF MICROSCOPY-OXFORD, 1994, 173 :39-51
[6]  
BURMEISTER JS, 1998, IN PRESS BIOMATERIAL
[7]  
BURMEISTER JS, 1996, J BIOMED MATER RES, V30, P12
[8]   FOCAL ADHESIONS - TRANSMEMBRANE JUNCTIONS BETWEEN THE EXTRACELLULAR-MATRIX AND THE CYTOSKELETON [J].
BURRIDGE, K ;
FATH, K ;
KELLY, T ;
NUCKOLLS, G ;
TURNER, C .
ANNUAL REVIEW OF CELL BIOLOGY, 1988, 4 :487-525
[9]   CORRECTION OF CELLULAR AUTOFLUORESCENCE IN FLOW-CYTOMETRY BY MATHEMATICAL-MODELING OF CELLULAR FLUORESCENCE [J].
CORSETTI, JP ;
SOTIRCHOS, SV ;
COX, C ;
COWLES, JW ;
LEARY, JF ;
BLUMBURG, N .
CYTOMETRY, 1988, 9 (06) :539-547
[10]  
EDMINSTON PL, 1992, APPL SPECTROSC, V47, P250