Purpose: This study aimed to investigate the variation of MFGM proteins in colostrum of Hu sheep under different dietary crude protein levels. Experimental design: Collecting the colostrum from Hu sheep fed three different levels of dietary crude protein levels. Plasma membrane proteins were enriched, digested, labeled using an iTRAQ kit, and identified by LC-MS. Differentially expressed proteins were analyzed using bioinformatics technology. Results: Pair-wise comparisons among three treatments identified 286 differentially expressed MFGM proteins. Bioinformatics analysis suggested that the three most abundant differentially expressed MFGM proteins were signaling, disulfide-bonded, and secreted proteins.