Protein expression levels of membrane proteins in brain capillaries isolated from C57BL/6 WT and hMDR1 mice.

Brain capillary was isolated from the pooled frozen brains of 10 mice by means of nylon mesh method. Brain capillary passing through an 85 μm nylon mesh and retained on a 20 μm nylon mesh was collected, and digested with lysyl endopeptidase and trypsin. The digest was subjected to LC-MS/MS analysis with internal standard peptides. The each target peptide was monitored with four different SRM/MRM transitions. The preparation of pooled brain capillary from 10 mice was conducted only one time. For the pooled brain capillary, the digestion followed by LC-MS/MS analysis was repeated three times. The protein expression level of target molecule was determined as an average and variability (mean ± S.E.M.) of the quantitative values obtained from different SRM/MRM transitions in three analyses. Therefore, the S.E.M. represents the variability of protein expression level determined in different MS/MS transitions in three analyses, but does not represent the inter-individual variability. U.L.Q., under the limit of quantification. *p < 0.01, significantly different from the C57BL/6 WT mice. a measured by using a peptide probe set specific for human MDR1. b measured by using a peptide probe set specific for mouse mdr1a. c measured by using a peptide probe set common for human MDR1 and mouse mdr1a. Protein expression levels of membrane proteins in brain capillaries isolated from C57BL/6 WT and hMDR1 mice.