Homogenous populations of cells make up individual tissues, yet how organisms achieve such homogeneity is unknown. Le et al. use the C. elegans intestine to reveal that an initiator of RNA silencing is segregated unequally between cells. Suppression of this inequality during early development achieves tissue homogeneity.

Intracellular detection of antigens at the single cell level by flow cytometry requires optimized buffers and methods for detection of a given specificity and the compartment probed. Using a new buffer system tightly controlled for low non-specific staining, increased resolution, and significantly lower cell loss, we have developed an optimized system for detection of intracellular proteins by flow cytometry. This system works across multiple tissues from human and mouse models offering a one-method system for reliable antibody cross-reactivity tests. The optimized system detects proteins within the nucleus, endoplasmic reticulum, outer mitochondrial membrane, and the Golgi complex, with focus on optimal resolution of DNA-binding transcription factors. Intracellular targets detected included; FoxP3, T-bet, RORγT, GATA-3, Satb1, Bcl6, Bcl2, pH2AX, Ki-67, Themis, Nanog, Sox1, Sox2, Oct 3/4, Pax-6, Nestin, and various cytokines. In addition, fluorescent antibodies to cell surface antigens for multi-color flow were also found to be compatible with the optimized buffer system. Further verification of the buffer was done by high throughput screening of human cell surface markers with the BD LyoplateTM on activated human T-regulatory cells (Treg). GARP positive and negative Treg subsets were screened for various markers and the results are discussed in the poster.