# Troubleshooting the Infinium HD FFPE QC Assay Protocol

The [Infinium HD FFPE QC Assay Protocol](https://support.illumina.com/downloads/infinium_hd_ffpe_qc_assay_insert_\(15020981_rev_c\).html) determines whether DNA samples are good candidates for the Infinium HD FFPE Restore Protocol.

With unexpected results, consider these troubleshooting tips:

* Confirm that samples are quantified using a fluorometric method (e.g., Qubit or PicoGreen).
  * See the **Preparation** section of the protocol.
* Confirm that the thermocycler settings are specific to the qPCR master mix used.
  * See Step 11 of the protocol:\
    \- 50C\* for 2 minutes and/or 95C\* for 10 minutes are included **\*if required by the Master Mix manufacturer**.
* In this protocol, Ct (threshhold cycle) is referred to as Cq (quantification cycle).
* Confirm that the qPCR run is set to perform the correct analysis.
  * See **Analyzing Data** step 4 of the protocol.
  * Analysis must subtract average Cq values of samples from average Cq values of the QCT\_ST sample.
  * Analysis must not quantify Cq values against a standard curve.
  * If needed, consult the qPCR system/software manufacturer for support on how to set up the run/analysis correctly.
* It is acceptable to have negative Delta Cq values for the samples. This result means that the DNA is more concentrated than the QCT\_ST standard sample.
  * Importantly, the data is acceptable if the the NTC (no template control) Cq is > 10 cycles after the QCT\_ST Cq values.
  * Samples with Delta Cq values < 5 (including negative Delta Cq values) are good candidates for restoration.

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