A simple spectrofluorometric assay to measure total intracellular magnesium by a hydroxyquinoline derivative.
The intracellular behaviour of diaza-18-crown-6 appended with two H-substituted hydroxyquinoline groups (DCHQ1) was investigated to explore its application as a new sensor for the evaluation of cell magnesium content and distribution. We used five cells lines characterised by different contents of magnesium and different intracellular membrane-defined compartments. The main result is the definition of the appropriate experimental conditions to quantitatively assess the total cell magnesium by fluorescence spectroscopy. We showed that disrupting cells by sonication, DCHQ1 was capable to assess total cell magnesium in all cell types examined, obtaining overlapping results with atomic absorption spectroscopy (AAS). This new analytical approach requires very small cell samples and a simple fluorimetric technique, and can be a valid alternative to AAS. The fluorescent properties of DCHQ1 in living cells are: (a) it consistently stains live cells, (b) it discriminates small variations of cell Mg contents, (c) cell staining is stable for at least 30 min. We also investigated the role of lipophilic environment on DCHQ1 fluorescence by mimicking cell membranes and described how the composition and structure of lipid vesicles affect Mg-DCHQ1 fluorescence. Thus, DCHQ1 may offer important information also on magnesium distribution in living cells, providing a novel strategy to map the intracellular compartmentalization of this cation.
Dipartimento di Biochimica G. Moruzzi, Università degli Studi di Bologna, Via S. Donato 19/2, 40127, Bologna, Italy. Giovanna.Farruggia@unibo.it
Giovanna
G
Farruggia
Stefano
S
Iotti
Luca
L
Prodi
Nelsi
N
Zaccheroni
Marco
M
Montalti
Paul B
PB
Savage
Giulia
G
Andreani
Valentina
V
Trapani
Federica I
FI
Wolf
D000818:Animals; D019556:COS Cells; D002460:Cell Line; D002522:Chlorocebus aethiops; D047248:Crown Compounds / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry; D018922:HL-60 Cells; D006801:Humans; D006912:Hydroxyquinolines / Q000737:chemistry*; D042541:Intracellular Space / Q000737:chemistry*; D008274:Magnesium / Q000032:analysis* / Q000737:chemistry; D051379:Mice; D051381:Rats; D012680:Sensitivity and Specificity; D013050:Spectrometry, Fluorescence / Q000379:methods*
D047248:Crown Compounds; D005456:Fluorescent Dyes; D006912:Hydroxyquinolines; D008274:Magnesium