Ambient RNA Correction

Subtract a caller-estimated ambient RNA component from cell counts.

Description

An ambient RNA estimate distributes a supplied molecule amount over a normalized background profile.

An ambient RNA estimate distributes a supplied molecule amount over a normalized background profile. Subtraction is capped at each feature's observed count.^[1]

Subtract a caller-estimated ambient RNA component from cell counts.

When to use Ambient RNA Correction

Use this tool to subtract a caller-estimated ambient RNA component from cell counts.

Prepare the upstream counts, expression values, sequences or record lists requested by this tool. Keep corresponding features in the same order and confirm count units, transformations and caller-selected criteria.

How to use Ambient RNA Correction and understand its results

Prepare the upstream counts, expression values, sequences or record lists requested by this tool. Keep corresponding features in the same order and confirm count units, transformations and caller-selected criteria.
Enter observedCounts, ambientProfile, ambientMolecules using the field table and calculation rules, then run the tool.
Read correctedCounts, removedMolecules. The explanation below defines the units, signs, ordering and any flags or placeholder values.
If the tool reports an input issue, correct the values and run it again. Apply the stated assumptions when interpreting a successfully returned result.

Input field	Meaning	Value type; see rules below	Requirement
observedCounts	Observed counts	list of number	Required
ambientProfile	Ambient profile	list of number	Required
ambientMolecules	Ambient molecules	number	Required

Enter aligned nonempty nonnegative counts/profile values and nonnegative ambient amount. Profile total must be positive even when ambient amount is zero. Normalize the profile, subtract and floor at zero. Return corrected values and total actually removed.

corrected_i=max(0,observed_i−ambientMolecules×profile_i/Σprofile)

Units, indexing, boundary conditions and exceptional cases are defined in the calculation rules above.

Output field	Meaning	Value type
correctedCounts	Corrected counts	list of number
removedMolecules	Removed molecules	number

json

{
  "input": {
    "observedCounts": [
      100,
      50
    ],
    "ambientProfile": [
      0.5,
      0.5
    ],
    "ambientMolecules": 20
  },
  "output": {
    "correctedCounts": [
      90,
      40
    ],
    "removedMolecules": 20
  }
}

Limitations and assumptions

Removal can be smaller than the requested amount due to clipping and corrected counts may be fractional. No ambient component is estimated, integer allocation performed or full contamination model fitted.

Numeric inputs must be finite. Follow the calculation-specific rules for overflow, underflow and exceptional cases. Lists are limited to 100,000 entries; the rules above may require smaller limits. Missing required fields, extra fields, wrong value types and violations of the calculation rules are input errors.

Alternative or Complementary methods

Use Empty Droplet Filter: Classify droplets by a caller-supplied minimum UMI count. For a complementary task, use Counts Per Million: Convert raw feature counts into counts per million mapped reads.

References

Ambient RNA Correction — tool.io

Similar or alternative tools

Empty Droplet Filter
Classify droplets by a caller-supplied minimum UMI count.
Counts Per Million
Convert raw feature counts into counts per million mapped reads.