bead-count-threshold-filtering

bead-count-threshold-filtering

Summary

Remove low-quality probes from methylation array data by filtering out probes with fewer than 3 beads in at least 5% of samples. This quality control step eliminates unreliable measurements before downstream analysis on HumanMethylation450 or EPIC arrays.

When to use

Apply this filter after loading raw .idat files or beta-valued matrices from HumanMethylation450 or EPIC methylation arrays when you need to remove probes with insufficient bead counts that may introduce measurement noise or bias into downstream differential methylation or enrichment analyses.

When NOT to use

• Input probes have already been filtered for bead count or quality — applying champ.filter() again risks over-filtering and loss of biological signal.
• Analysis requires probes at the boundaries of technical reliability for hypothesis-driven validation — the threshold may exclude important but marginal probes.
• Bead count data is not available or has been discarded during preprocessing — the filter cannot be applied without this information.

Inputs

• HumanMethylation450 or EPIC array intensity data (in RGChannelSet or MethylSet format)
• Detection p-value matrix (optional, for sequential filtering context)
• Bead count matrix (automatically generated from .idat files or provided separately)

Outputs

• Filtered probe matrix with low-bead-count probes removed
• Pre- and post-filter probe count comparison
• Quality control report documenting number of probes retained and removed
• Bead count distribution plots (before and after filtering)

How to apply

Use ChAMP's champ.filter() function with default parameters, which applies bead-count filtering as the second successive quality control step (after detection p-value filtering). The filter removes any probe where fewer than 3 beads are detected in at least 5% of samples in the dataset. This threshold is based on the Illumina bead array technical design, where probes with fewer than 3 beads per sample are considered unreliable. Execute the filter on the full probe set, then compare pre- and post-filter probe counts and bead count distributions to verify that low-bead probes have been removed and that the majority of probes (expected >95% retention) remain for analysis.