Histone post-translational modifications (PTMs) create a powerful regulatory mechanism for maintaining chromosomal integrity in cells.
Histone acetylation and methylation, the most widely studied
histone PTMs, act in concert with
chromatin-associated
proteins to control access to genetic information during transcription. Alterations in cellular
histone PTMs have been linked to disease states and have crucial
biomarker and therapeutic potential. Traditional bottom-up mass spectrometry of
histones requires large numbers of cells, typically one million or more. However, for some cell subtype-specific studies, it is difficult or impossible to obtain such large numbers of cells and quantification of rare
histone PTMs is often unachievable. An established targeted LC-MS/MS method was used to quantify the abundance of
histone PTMs from cell lines and primary human specimens. Sample preparation was modified by omitting nuclear isolation and reducing the rounds of
histone derivatization to improve detection of
histone peptides down to 1,000 cells. In the current study, we developed and validated a quantitative LC-MS/MS approach tailored for a targeted
histone assay of 75
histone peptides with as few as 10,000 cells. Furthermore, we were able to detect and quantify 61
histone peptides from just 1,000 primary human stem cells. Detection of 37
histone peptides was possible from 1,000
acute myeloid leukemia patient cells. We anticipate that this revised method can be used in many applications where achieving large cell numbers is challenging, including rare human cell populations.