ac8b03692_si_001.xlsx (54.18 kB)
Download fileIntegrated Proteome Analysis Device for Fast Single-Cell Protein Profiling
dataset
posted on 2018-10-30, 00:00 authored by Xi Shao, Xuantang Wang, Sheng Guan, Haizhu Lin, Guoquan Yan, Mingxia Gao, Chunhui Deng, Xiangmin ZhangIn
our previous work, we have demonstrated an integrated proteome
analysis device (iPAD-100) to analyze proteomes from 100 cells. In this work, for the first time, a novel integrated
device for single-cell analysis (iPAD-1) was developed to profile
proteins in a single cell within 1 h. In the iPAD-1, a selected single
cell was directly sucked into a 22 μm i.d. capillary. Then the
cell lysis and protein digestion were simultaneously accomplished
in the capillary in a 2 nL volume, which could prevent protein loss
and excessive dilution. Digestion was accelerated by using elevated
temperature with ultrasonication. The whole time of cell treatment
was 30 min. After that, single-cell digest peptides were transferred
into an LC column directly through a true zero dead volume union,
to minimize protein transfer loss. A homemade 22 μm i.d. nano-LC
packing column with 3 μm i.d. ESI tip was used in the device
to achieve ultrasensitive detection. A 30 min elution program was
applied to analysis of the single-cell proteome. Therefore, the total
time needed for a single-cell analysis was only 1 h. In an analysis
of 10 single HeLa cells, a maximum of 328 proteins were identified
in one cell by using an Orbitrap Fusion Tribrid MS instrument, and
the detection limit was estimated at around 1.7–170 zmol. Such
a sensitivity of the iPAD-1 was ∼120-fold higher than that
of our previously developed iPAD-100 system. Prominent cellular heterogeneity in protein expressive profiling
was observed. Furthermore, we roughly estimated the phases of the
cell cycle of tested HeLa cells by the amount of core histone proteins.