Positive and negative ion modes: using the same sample, changing the ‘polarity’ reveals the other half of the molecule

Mass spectrometry acquisition is typically divided into two modes: positive ion mode (+) and negative ion mode (−). In positive mode, the instrument prioritises the detection of positively charged ions, typically such as [M+H]⁺ and [M+Na]⁺, formed when molecules combine with H⁺, Na⁺ or K⁺; in negative ion mode, the instrument prioritises the detection of negatively charged ions, typically those formed when a molecule loses H⁺ or gains Cl⁻, such as [M−H]⁻ and [M+Cl]⁻. Switching between positive and negative modes essentially involves changing which type of charge the instrument is ‘willing to collect’.

Different molecules are inherently suited to different modes. Molecules containing basic groups (such as amino groups) are easily protonated, resulting in strong signals in positive mode; molecules containing acidic groups (such as carboxylic acids and phosphates) are easily deprotonated, resulting in strong signals in the negative mode. Consequently, the spectra obtained from the same sample in these two modes are often complementary—each illuminates half of the molecules, and only when combined do they provide a true picture of the sample. Running a single mode alone is likely to result in the systematic omission of the other class of compounds.

This complementarity is particularly valuable in mass spectrometry imaging. Take MSI DPI imaging of mouse brain sections as an example: in positive ion mode, the neurotransmitter GABA, creatine, glutamine, glutamate, adenosine, as well as cholesterol and more than a dozen neutral GalCer lipids were newly detected; whilst in negative ion mode, the analysis covered phosphatidylethanolamine (PE), acetalated phospholipid PE-O, phosphatidylserine (PS), phosphatidylinositol (PI), sulpholipids (ST) and others, with PE-O/PE signals increased by 2–4-fold. The simultaneous imaging of neurotransmitters and various lipids holds direct significance for research in neuroscience, pharmacology and neurochemistry.

The same applies to research on melanocytic naevi: positive mode primarily visualises choline, phosphocoline, S1P, cholesterol, phosphatidylcholine (PC) and sphingomyelin (SM), whilst negative mode visualises fatty acids (FA), PE, phosphatidic acid (PA), PI and others; only by combining the two modes can the lipid differences between the naevus area and normal tissue be fully characterised. Neo Source INSTRUMENT’s MSI DPI and MSI LDPI both support switching between positive and negative ion modes and can be retrofitted onto mass spectrometers from Agilent, AB SCIEX and Thermo.