Nano-scale liquid chromatography-mass spectrometry(nanoLC-MS)has been a routine tool in proteomic research,however,the analysis of trace sample(e.g.,cells from small tissue samples obtained using laser capture microdissection,stem cells from the blood)can still be problematic and the sensitivity should be further improved.In this work,20-μm-i.d.C18-functionalized amine-bridged hybrid monolith was prepared and used as the separation column with low backpressure,high resolution and low non-specific adsorption.Furthermore,integrated gold coated emitter with stable nanoelectrospray was prepared for coupling such narrow-bore separation column to MS.By such nanoLC-MS analysis,16 unique peptides could be identified from 5 pg BSA digests,and the sequence coverage of 35.26%was obtained.The peak width at half height is only 4 s and high peak capacity of 342 was achieved.Finally,the digests from Hela cells were used as sample and analyzed with high sensitivity.721 protein groups were identified from 10 ng samples in the single run and 66 protein groups from 2 ng samples.Besides,we also prepared 10-μm-i.d.open-tubular capillary columns coated with C18 polymer brushes by surface initiated atom transfer radical polymerization,followed by combination with MS.In a 50-min run,~1000 proteins could be identified from 10 ng digests from Hela cells.Totally,3035 peptides corresponding to 1327 proteins were identified in three runs.It is indicated that these ultrasensitive methods have great potential to solve the bottleneck problems of proteome analysis in trace sample,and provide novel technical support to improve the qualitative and quantitative analytical capacity for proteomic study.