Capillary electrophoresis-electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry for direct analysis of cellular proteins.
The combination of capillary electrophoresis (CE) with electrospray ionization (ESI) mass spectrometry has proven to be broadly applicable to a wide range of biologically important compounds. When combined with Fourier transform ion cyclotron resonance (FTICR) mass spectrometry, the combined method, in addition to high-resolution separations, affords high-resolution precision mass measurements for analytes separated from complex mixtures. Direct chemical analysis of single cells has received considerable attention in recent years; the single cell approach provides a major step toward answering important questions in the field of cellular biochemistry. In this work we present preliminary results which demonstrate the feasibility of using the CE-ESI-FTICR combination as a high-performance detection scheme for the analysis of cellular proteins acquired directly from small populations (i.e., 5-10) of intact living cells. The human erythrocyte was chosen as a model system owing to its availability, relatively homogeneous composition, and thorough documentation of contents by previous researchers. In this work we demonstrate the on-line acquisition of high-resolution mass spectra (average resolution > or = 45,000 fwhm) of both the alpha and the beta chains of hemoglobin acquired from the injection of 10 human erythrocytes (corresponding to 4.5 fmol of hemoglobin). Given the extremely small volume of the human erythrocyte (typically 87 fl/cell), the techniques implemented here should also be adaptable to the study of larger mammalian cell systems.
Hofstadler
S A
SA
Chemical Sciences Department, Pacific Northwest Laboratory, Richland, Washington 99352, USA.
Swanek
F D
FD
Gale
D C
DC
Ewing
A G
AG
Smith
R D
RD
D000328:Adult; D017357:Cyclotrons; D004586:Electrophoresis / Q000379:methods*; D004912:Erythrocytes / Q000737:chemistry; D005583:Fourier Analysis; D006454:Hemoglobins / Q000032:analysis; D006801:Humans; D008297:Male; D013058:Mass Spectrometry / Q000379:methods*; D011506:Proteins / Q000032:analysis*
D006454:Hemoglobins; D011506:Proteins