Neuropeptides are low abundance signaling molecules that modulate almost every physiological process, and dysregulation of neuropeptides is implicated in disease pathology. Mass spectrometry (MS) imaging is becoming increasingly useful for studying neuropeptides as new sample preparation methods for improving neuropeptide detection are developed. In particular, proper tissue washes prior to MS imaging have shown to be quick and effective strategies for increasing the number of detectable neuropeptides. Treating tissues with solvents could result in either gain or loss of detection of analytes, and characterization of these wash effects is important for studies targeting sub-classes of neuropeptides. In this communication, we apply aqueous tissue washes that contain sodium phosphate salts, including 10% neutral buffered formalin (NBF), on crustacean brain tissues. Our optimized method resulted in complementary identification of neuropeptides between washed and unwashed tissues, indicating that our wash protocol may be used to increase total neuropeptide identifications. Finally, we show that identical neuropeptides were detected between tissues treated with 10% NBF and an aqueous 1% w/v sodium phosphate solution (composition of 10% NBF without formaldehyde), suggesting that utilizing a salt solution wash affects neuropeptide detection and formaldehyde does not affect neuropeptide detection when our wash protocol is performed.
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The datasets generated during and/or analyzed during the current study are available in the MassIVE repository: https://doi.org/10.25345/C5DQ8H. MassIVE DOI: MSV000085824.
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This work was supported in part by a National Science Foundation grant (CHE-1710140) and the National Institutes of Health (NIH) through grants 1R01DK071801 and R56 MH110215. N.Q.V. would like to thank the National Heart, Lung, and Blood Institute of the NIH under the Training Program in Translational Cardiovascular Science (T32 HL007936-20) for funding. A.R.B. was funded through the NIH in the form of a General Medical Sciences NRSA Fellowship 1F31GM119365. J.J would like to acknowledge the University of Wisconsin Carbone Cancer Center (233-AAC9675) and the UWCCC Pancreatic Cancer Taskforce for Funding. L.L. acknowledges a Vilas Distinguished Achievement Professorship and Charles Melbourne Johnson Professorship with funding provided by the Wisconsin Alumni Research Foundation and University of Wisconsin-Madison School of Pharmacy. The MALDI-Orbitrap instrument was purchased through the support of an NIH shared instrument grant (S10RR029531) and Office of the Vice Chancellor for Research and Graduate Education at the University of Wisconsin−Madison.
Conflict of interest
The authors declare that they have no conflicts of interest.
In-house crustacean neuropeptide database is available from the corresponding author upon reasonable request.
Published in the topical collection Mass Spectrometry Imaging 2.0 with guest editors Shane R. Ellis and Tiffany Siegel Porta.
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Vu, N.Q., Buchberger, A.R., Johnson, J. et al. Complementary neuropeptide detection in crustacean brain by mass spectrometry imaging using formalin and alternative aqueous tissue washes. Anal Bioanal Chem 413, 2665–2673 (2021). https://doi.org/10.1007/s00216-020-03073-x
- Mass spectrometry imaging