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ProteinSimple

Milo Single Cell Western

milo
milo_chip
Milo™ lets you identify cellular subpopulations that aren't visible in traditional Western Blots, so you can study distinct cell types in your heterogeneous sample. Have a protein target you can't measure with flow cytometry? Try Milo! He uses off-the-shelf, Western-validated antibodies and can even measure intracellular proteins, transcription factors, phosphorylated proteins, or targets that don't have flow antibodies. Already doing single-cell RNA studies? Milo gives you direct protein validation of your single-cell RNA targets, so you can be way more confident that your gene expression results are real.

ÁRAJÁNLATOT, INFORMÁCIÓT KÉREK!

HOW IT WORKS

Milo uses scWest™ chips to get your single-cell Western results in about four hours! All you do is load your cell suspension and the scWest chip partitions the cells and captures ~1,000 individual cells for you. Milo then does a fast, 1 minute SDS-PAGE separation on each single-cell lysate on-chip. Then just use your favorite conventional Western antibodies to probe the chip for up to 4 targets per cell simultaneously.

FIGURE 1.  scWest chip and an array of Single-Cell Westerns probed for two protein targets in each single-cell lysate.

SUBPOPULATION AND PROTEIN ISOFORM IDENTIFICATION

Milo measures protein expression in ~1,000 individual cells. That means you can identify which cells in your sample express each protein target. With that info, you'll be able to identify subpopulations of cells that can't be measured with a conventional, population-level Western Blot. As an added bonus, Milo also resolves protein isoforms that can't be distinguished with flow cytometry. Milo gives you info about your cells that you just can't get any other way!

 

FIGURE 2.  One antibody binds to two protein variants (Exon 1 and Exon 2) expressed in HEK293 cells. Milo reveals three subpopulations of cells: cells expressing only a large molecular weight variant (Exon 1), only a small molecular weight variant (Exon 2) or both. In contrast, conventional Western blotting can only detect average expression in the mixed population.

PROTEIN EXPRESSION HETEROGENEITY

Now you can easily interrogate what cell subtypes are contributing to your bulk sample behavior. So if you're wondering if all your cells are expressing your target equally or if you've got some high expressers and some low expressers—Milo can help!

FIGURE 3.  Milo reveals low and high KRT8 expressing cells within a human primary breast epithelial sample.

Műszaki specifikációk

 

Requirements &compatibility.

  • Sample type: Suspension containing >10,000 cells
  • Cell diameter: 7–25µm
  • Cell type:Mammalian cells; globular in suspension and unfixed
  • Antibody requirement: Standard, commercial unlabeled primaries and fluorescent secondaries
  • Other equipment needed: Open-format fluorescence microarray scanner capable of 10µm resolution

 

Performance &specifications.

  • Typical cell dilutions yield capture and analysis of 1,000–2,000 cells per scWestchip
  • Molecular weight range: 15–175kDa
  • MW resolution: 10% differences in distinct spectral channels, as low as 30% differences in same spectral channel
  • Typical target multiplexing: Up to four proteins per cell by spectral and size-basedmultiplexing
  • Workflow time: 4–6 hours

 

 

Applikációk

SUBPOPULATION AND PROTEIN ISOFORM IDENTIFICATION

Milo measures protein expression in ~1,000 individual cells. That means you can identify which cells in your sample express each protein target. With that info, you'll be able to identify subpopulations of cells that can't be measured with a conventional, population-level Western Blot. As an added bonus, Milo also resolves protein isoforms that can't be distinguished with flow cytometry. Milo gives you info about your cells that you just can't get any other way!

 

FIGURE 2.  One antibody binds to two protein variants (Exon 1 and Exon 2) expressed in HEK293 cells. Milo reveals three subpopulations of cells: cells expressing only a large molecular weight variant (Exon 1), only a small molecular weight variant (Exon 2) or both. In contrast, conventional Western blotting can only detect average expression in the mixed population.

PROTEIN EXPRESSION HETEROGENEITY

Now you can easily interrogate what cell subtypes are contributing to your bulk sample behavior. So if you're wondering if all your cells are expressing your target equally or if you've got some high expressers and some low expressers—Milo can help!

FIGURE 3.  Milo reveals low and high KRT8 expressing cells within a human primary breast epithelial sample.

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