qHCR Flow Cytometry

qHCR flow cytometry enables RNA relative quantitation for high-throughput expression profiling of mammalian and bacterial cells without the need to engineer reporter lines.

 

Analog Quantitative Signal

qHCR signal is analog in the form of single-cell fluorescence intensities that scale approximately linearly with the number of RNA target molecules per cell. 

Quantitative RNA flow cytometry

Mammalian Cells (HEK)

Redundant 2-Channel Detection

To illustrate quantitative accuracy and precision for single-cell measurements with RNA flow cytometry, we detect a target mRNA using two probe sets that trigger different HCR amplifiers carrying spectrally distinct fluorophores.  

Bacterial Cells (E. coli)

High Accuracy and Precision

In the resulting scatter plot of fluorescence single-cell intensities, high accuracy corresponds to linearity with zero intercept, and high precision corresponds to tight scatter around the line. 

Straightforward Multiplexing

Orthogonal HCR amplifiers enable 1-step signal amplification for up to 5 target RNAs simultaneously. Straightforward multiplexing is achieved using the same 1-day protocol independent of the number of target RNAs. Learn more

Custom Probe Set Design

Custom probe set design is available for any target mRNA in any organism across the tree of life. Learn more

Automatic Background Suppression

Using HCR v3.0 probes and amplifiers, qHCR flow cytometry benefits from automatic background suppression throughout the protocol, dramatically enhancing performance and ease-of-use. Learn more

Precision Increases with Probe Set Size

Automatic background suppression ensures that precision increases with probe set size. As a result, for qHCR flow cytometry, we recommend using 30 or more split-initiator probe pairs per target RNA (maximize probe set size given target length). 

Signal-to-Background Increases with Amplification Time 

Signal increases with amplification time and background does not, so for qHCR flow cytometry, we recommend overnight amplification to grow long amplification polymers and enhance signal-to-background. 

qHCR Flow Cytometry
 Analog RNA relative quantitation

✓ High accuracy and precision

✓ High-throughput single-cell assay

✓ Automatic background suppression

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