Summary
Matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) can provide rapid, sensitive determinations of lipids from small tissue samples in both single determinations and automated high-throughput assays. MALDI-TOF MS is a sensitive, high-throughput technique for the determination of lipids such as the phosphoinositides, PtdIns (phosphatidylinositol), PIP (phosphatidylinositol-4-phosphate, and PIP2 (phosphatidylinositol-4,5-bisphosphate), but in crude extracts the signals are weak or not observed due in large part to ion suppression by phosphatidylcholine and other cationic lipids. A rapid separation step using a small column of a strong cation exchange (SCX) gel can be utilized easily and effectively to adsorb or capture cationic lipids from chloroform/methanol lipid extracts and provide substantially improved signals for the phosphoinositides. In this review, we describe the use of fractionation of a crude lipid extracts using cation exchange columns in conjunction with MALDI-TOF MS and appropriate internal standards to quantify the levels of phosphoinositides in small mammalian brain samples.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Han, X. and Gross, R. W. (2005) Shotgun lipidomics: electrospray ionization mass spectrometric analysis and quantitation of cellular lipidomes directly from crude extracts of biological samples. Mass Spectrom. Rev. 24, 367–412.
Han, X. and Cheng, H. (2005) Characterization and direct quantitation of cerebroside molecular species from lipid extracts by shotgun lipidomics. J. Lipid Res. 46, 163–175.
Schiller, J., Suss, R., Arnhold, J., Fuchs, B., Lessig, J., Muller, M., Petkovic, M., Spalteholz, H., Zschornig, O., and Arnold, K. (2004) Matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry in lipid and phospholipid research. Prog. Lipid Res. 43, 449–488.
Johanson, R. A., Buccafusca, R., Quong, J. N., Shaw, M. A., and Berry, G. T. (2007) Phosphatidylcholine removal from brain lipid extracts expands lipid detection and enhances phosphoinositide quantification by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. Anal. Biochem. 362, 155–167.
Milne, S., Ivanova, P., Forrester, J., and Alex, Brown, H. (2006) Lipidomics: an analysis of cellular lipids by ESI-MS. Methods 39, 92–103.
Petkovic, M., Schiller, J., Muller, M., Benard, S., Reichl, S., Arnold, K., and Arnhold, J. (2001) Detection of individual phospholipids in lipid mixtures by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry: phosphatidylcholine prevents the detection of further species. Anal. Biochem. 289, 202–216.
Schiller, J., Arnhold, J., Benard, S., Muller, M., Reichl, S., and Arnold, K. (1999) Lipid analysis by matrix-assisted laser desorption and ionization mass spectrometry: A methodological approach. Anal. Biochem. 267, 46–56.
Muller, M., Schiller, J., Petkovic, M., Oehrl, W., Heinze, R., Wetzker, R., Arnold, K., and Arnhold, J. (2001) Limits for the detection of (poly-)phosphoinositides by matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF MS). Chem. Phys. Lipids 110, 151–164.
Schiller, J., Suss, R., Fuchs, B., Muller, M., Zschornig, O., and Arnold, K. (2003) Combined application of TLC and matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF MS) to phospholipid analysis of brain. Chromatographia 57, S297–S302.
Schacht, J. (1981) Extraction and purification of polyphosphoinositides. Methods Enzymol. 72, 626–631.
Arthur, G. and Sheltawy, A. (1980) The presence of lysophosphatidylcholine in chromaffin granules. Biochem. J. 191, 523–532.
Christie, W. W. (1993) (Ed.), Advances in Lipid Methodology – Two, Oily Press, Dundee, 1993, pp. 195–213. (available online at http://www.lipidlibrary.co.uk/topics/extract2/index.htm)
Hauser, G., Eichberg, J., and Gonzalez-Sastre, F. (1971) Regional distribution of polyphosphoinositides in rat brain. Biochim. Biophys. Acta 248, 87–95.
Hauser, G. and Eichberg, J. (1973) Improved conditions for the preservation and extraction of polyphosphoinositides. Biochim. Biophys. Acta 326, 201–209.
Nishihara, M. and Keenan, R. W. (1985) Inositol phospholipid levels of rat forebrain obtained by freeze-blowing method. Biochim. Biophys. Acta 835, 415–418.
Bligh, E. G. and Dyer, W. J. (1959) A rapid method for total lipid extraction and purification. Can. J. Biochem. Physiol. 37, 911–917.
Hendrickson, H. S. and Ballou, C. E. (1964) Ion exchange chromatography of intact brain phosphoinositides on diethylaminoethyl cellulose by gradient salt elution in a mixed solvent system. J. Biol. Chem. 239, 1369–1373.
Kiselev, G. V. (1982) Preparative isolation of polyphosphoinositide fractions from ox brain. Biochim. Biophys. Acta 712, 719–721.
Low, M. G. (1990) Purification of Phosphatidylinositol 4-Phosphate and Phosphatidylinositol 4,5-Bisphosphate by Column Chromatography. In: Methods in Inositide Research, (Irvine, R. F. ed.) Raven Press, New York, pp.145–151.
Baldwin, M. A. (2005) Mass spectrometers for the analysis of biomolecules. Methods Enzymol. 402, 3–48.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2009 Humana Press, a part of Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Johanson, R.A., Berry, G.T. (2009). Brain Phosphoinositide Extraction, Fractionation, and Analysis by MALDI-TOF MS. In: Armstrong, D. (eds) Lipidomics. Methods in Molecular Biology, vol 579. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60761-322-0_9
Download citation
DOI: https://doi.org/10.1007/978-1-60761-322-0_9
Published:
Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-60761-321-3
Online ISBN: 978-1-60761-322-0
eBook Packages: Springer Protocols