Gradient, high-resolution, magic angle spinning 1H nuclear magnetic resonance spectroscopy of intact cells.
The application of new gradient, high-resolution, magic angle spinning (MAS) 1H nuclear magnetic resonance (NMR) spectroscopy to the study of intact undifferentiated and differentiated NIH 3T3 F442A cells demonstrated improved spectral resolution and sensitivity compared with static studies. MAS of cells permits the detection and quantitation of many cellular metabolites that are not clearly resolved in nonspinning measurements and provides an improved visibility of phospholipids. Gradient, MAS enables the use of diffusion weighting for compartment assignment and the determination of mobility for many metabolites which are incompletely resolved using static techniques. The smaller, undifferentiated preadipocytes show no microscopic evidence of cell lysis after 2 h of MAS at 3.5 kHz and 82% of these cells remain viable by trypan blue exclusion. In contrast, 15-19% of the larger, lipid-laden differentiated adipocytes were found to suffer some degree of cell lysis with MAS. This new method is an attractive alternative to either nonspinning perfusion or extraction techniques for NMR studies of cells.
Weybright
P
P
Department of Surgery, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
Millis
K
K
Campbell
N
N
Cory
D G
DG
Singer
S
S
D016475:3T3 Cells; D017667:Adipocytes / Q000166:cytology / Q000378:metabolism*; D000818:Animals; D002452:Cell Count; D002454:Cell Differentiation; D002460:Cell Line; D002462:Cell Membrane / Q000648:ultrastructure; D002470:Cell Survival; D002794:Choline / Q000032:analysis; D004396:Coloring Agents; D004058:Diffusion; D006838:Hydrocarbons; D006859:Hydrogen; D007294:Inositol / Q000032:analysis; D050356:Lipid Metabolism; D008055:Lipids / Q000032:analysis; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D008563:Membrane Lipids / Q000032:analysis; D008697:Methane / Q000031:analogs & derivatives / Q000032:analysis; D051379:Mice; D010713:Phosphatidylcholines / Q000032:analysis; D010743:Phospholipids / Q000032:analysis / Q000378:metabolism; D010767:Phosphorylcholine / Q000032:analysis; D014280:Triglycerides / Q000032:analysis; D014343:Trypan Blue
D004396:Coloring Agents; D006838:Hydrocarbons; D008055:Lipids; D008563:Membrane Lipids; D010713:Phosphatidylcholines; D010743:Phospholipids; D014280:Triglycerides; D010767:Phosphorylcholine; C030011:carbene; D007294:Inositol; D006859:Hydrogen; D014343:Trypan Blue; D002794:Choline; D008697:Methane