FIGURE 2.
RP-HPLC followed by HILIC allows a >104 dynamic range for detection of H4 forms. A, fraction 41 contains ∼0.07% of the total observed histone H4 area in the HILIC chromatogram (Fig. 1). B, FTMS of fraction 41 from A shows the presence of at least three different intact masses. C, ECD of the isolated mass 11387 Da in B identifies at least two isomeric forms present at ∼50% each. Using the MS and chromatographic signal levels, a dynamic range of ∼7 × 104 is possible, translating to the ability to detect a H4 form present in ∼500 nucleosomes of the ∼32 million total per cell. S/N, signal-to-noise ratio.