HCR Northern Blot Technology

Multiplexed quantitative RNA blotting

HCR probe sets, amplifiers, and buffers enable multiplexed quantification of RNA target size and abundance with automatic background suppression throughout the protocol.

Quantitative Signal

qHCR signal is quantitative in the form of band intensities that scale approximately linearly with target abundance.

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HCR Northern Blotting: How It Works

Detection Stage

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Amplification Stage

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Detection Stage

An HCR probe set comprises multiple probe pairs that hybridize to different subsequences along the target. HCR initiator i1 is split between each pair of probes such that only probe pairs that hybridize specifically to the target RNA colocalize the full HCR initiator i1.

Amplification Stage

An HCR amplifier comprises metastable fluorescent HCR hairpins h1 and h2. Specifically bound probe pairs that colocalize full HCR initiator i1 trigger growth of a tethered fluorescent HCR amplification polymer. 

Automatic Background Suppression

HCR probes and amplifiers combine to provide automatic background suppression throughout the protocol: probes that bind non-specifically do not colocalize full HCR initiator i1 and do not trigger HCR; hairpins that bind non-specifically are kinetically trapped and do not trigger HCR.

RNA Relative Quantitation via qHCR Northern Blot

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Relative Quantitation

For a given target, relative quantitation is performed by comparing band intensities between lanes. 

3-Plex RNA Relative Quantitation

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High Accuracy and Precision

Standard curves demonstrate high accuracy (linearity with zero intercept) and precision (tight scatter around the line). 

RNA Absolute Quantitation via qHCR Northern Blot

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Absolute Quantitation

Absolute quantitation is performed by comparing samples to a standard curve generated using RNA targets of known abundance.

Absolute miRNA Quantitation for HEK 293T and HeLa Cells

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High Accuracy and Precision

Standard curves demonstrate high accuracy (linearity with zero intercept) and precision (tight scatter around the line). 

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Simple Robust Protocols

qHCR northern blot protocols are simple, robust, and enzyme-free, requiring only 2 stages independent of the number of target RNAs.

Automatic Background Suppression

HCR v3.0 reagents provide automatic background suppression throughout the protocol, ensuring that even if probes or hairpins bind non-specifically in the sample they will not generate amplified background, dramatically enhancing performance and ease-of-use.

Straightforward Multiplexing

HCR enables straightforward multiplexing using 1-step quantitative signal amplification for all target RNAs simultaneously.

Custom Probe Design

Free custom probe design is available for any target mRNA in any organism across the tree of life.

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Reagents will not generate amplified background even if they bind non-specifically within the sample

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The same 2-stage enzyme-free protocol is used independent of the number of target RNAs

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Custom probe set design for any target mRNA in any organism across the tree of life

qHCR Northern Blots

 Simultaneously quantify RNA target size and abundance

✓ Relative and absolute quantitation

✓ High accuracy and precision

✓ Automatic background suppression throughout the protocol