Breast cancer originates from mammary epithelial cells due to genetic alterations in mammary epithelial cells leading to subsequent loss of tissue homeostasis. After extensive research, scientists have divided breast epithelial cells into several different subpopulations, but still cannot fully understand epithelial cell heterogeneity and differentiation spectrum.
Scientists at the University of California, Irvine, published an article in the latest issue of Nature Communication, using single-cell mRNA sequencing (ScRNAseq) to analyze the transcriptome of 25,790 mammary epithelial cells from seven different breast hyperplasia patients. The use of non-segregation analysis revealed the presence of three different epithelial cell populations, one basal cells and the other, luminal epithelial cells , which are divided into hormone-secreting L1 and hormone-responsive L2-type cells. 
The authors found from single-cell sequencing data that KRT8 is more highly expressed than L1 in L2-type cells. 
To elucidate the heterogeneity of KRT8 protein expression, the authors selected ProteinSimple Milo single-cell western blot. Milo divides cells into three subpopulations: negative, low expression, and high expression. And can accurately calculate the percentage of each subpopulation of cells, that is, the number.
L2 was also characterized by higher levels of KRT8 than L1
To quantify protein expression in individual cells, we utilized a recently developed single-cell western blot application (ProteinSimple, Milo) , which performs electrophoretic separation of the protein content of about 2000 cells per chip and subsequently probed with fluorescently labeled antibodies.
Applying single-cell western blotting to luminal and basal cells isolated by FACS identified three cell states, ie KRT8-negative, -low, and -high , which illustrates the usefulness of single cell Western blotting as a quantitative validation tool downstream of scRNAseq analyses. 
Single cell gene sequencing + Milo single cell western blot combination can completely analyze cell heterogeneity from gene level and protein level, and explain key life medical problems such as tumorigenesis and stem cell development. 
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