Review author's response: pitfalls in isolating lipid rafts (original) (raw)

Nature Reviews Neuroscience volume 8, page 567 (2007)Cite this article

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The letter by Nothdurfter et al. regarding the methodological pitfalls associated with isolating lipid raft membranes is a welcome discussion about a topic that has evoked considerable debate in this field. We thank the authors for voicing their insightful comments and for providing experimental observations and suggestions to overcome some of the technical problems facing those investigating membrane microdomains. Isolation of detergent-resistant membranes remains a near-universal approach for obtaining lipid raft/caveolae membranes; however, it is important to emphasize that this experimental manipulation results in biochemical and operationally defined membrane microdomains, rather than iconic structures. It is worth reiterating that alternative fractionation techniques, which do not rely on detergent solubility, have addressed some of these problems1, and that these are preferred by some investigators. Both detergent and non-detergent membrane isolations result in defined membrane domains that are probably consistent for a given technique and tissue type. Thus, it is incumbent upon investigators to repeat their own methods exactly from experiment to experiment, and importantly, to clearly explain in their published work complete details of the method so that others can faithfully repeat the results. This should include outlining the details of concentration of detergent, volumes used and the protein-to-detergent ratios. Note, for example, that in Figure 1, Nothdurfter et al. add an internal standard and suggest that tissue types might be a reason for variability in lipid raft/caveloae determinations. However, it would be most informative to see these tissue types compared in experiments using the same detergent conditions and the same density matrix (for example, both using 1% TX-100 and both separated using discontinuous sucrose gradients). As we suggest in our review article2, studies involving the intricacies of signal transduction and lipid rafts should use several complementary approaches, and whenever possible, should use techniques that can be tested in intact cells3.

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Authors and Affiliations

  1. Departments of Physiology & Biophysics, University of Illinois at Chicago, College of Medicine, Illinois, USA
    John A. Allen & Mark M. Rasenick
  2. Psychiatry, and the Graduate Program in Neuroscience, University of Illinois at Chicago, College of Medicine, Illinois, USA
    Mark M. Rasenick

Authors

  1. John A. Allen
  2. Mark M. Rasenick

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Correspondence toMark M. Rasenick.

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Allen, J., Rasenick, M. Review author's response: pitfalls in isolating lipid rafts.Nat Rev Neurosci 8, 567 (2007). https://doi.org/10.1038/nrn2059-c2

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