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# Understanding loading and percent loading concentration on the NextSeq 1000/2000
**Background**
Illumina recommends using the %Loading Concentration run metric alongside the %Occupied metric to optimize loading concentrations on the NextSeq 1000/2000. This is because the NextSeq 1000/2000 uses a unique patterned flow cell that requires a new metric to help accurately determine proper loading.
The %Loading Concentration was added to RTA3 release for NextSeq 1000/2000 Control Software v1.3 and above.
**IMPORTANT NOTES**
* This metric relies on the diversity and complexity of reads to estimate the appropriate loading concentration.
* Libraries containing low diversity or low complexity (ie, amplicon libraries) could potentially impact the accuracy of the %Loading concentration, so %Loading Concentration should not be used for libraries that have low diversity or high proportion of similar reads.
* For situations where %Loading Concentration cannot be used, it is best to review a combination of Percent Pass Filter (%PF), Q30, %Occupied, and PhiX alignment as seen below.
**Identifying the %Loading Concentration for a** **run**
The %Loading concentration for a sequencing run can be found within the **PrimaryAnalysisMetrics.csv** file in the run folder, as well as in run metrics in BaseSpace. The PrimaryAnalysisMetrics.csv file is only available once the run completes.
**PrimaryAnalysisMetrics.csv from the on-instrument run folder:** /usr/local/illumina/runs/\[runfolder]
PrimaryAnalysisMetrics.csv file:\
**For runs in Basespace:** the PrimaryAnalysisMetrics.csv file will not be present in the run files, but the metric will be displayed in the metrics tab:\
**Alternatives to %Loading Concentration**
For low diversity or low complexity libraries, use acombination of Percent Pass Filter (%PF), %Occupancy, Q30, and PhiX alignment to help with loading concentration optimization.
* For over-loaded runs, expect to see low passing filter, high %Occupancy, reduced quality scores and lower than expected PhiX alignment.
* For under-clustered runs, expect to see low passing filter, low %Occupancy, high quality and higher than expected PhiX alignment.
After reviewing metrics, increase or decrease loading concentration in **100 pM** increments for titrations.
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| *For any feedback or questions regarding this article (Illumina Knowledge Article #3121), contact Illumina Technical Support* [*techsupport@illumina.com*](mailto:techsupport@illumina.com?subject=Question%2FFeedback%20Regarding%20Illumina%20Knowledge%20Article%20#000003121%20-%20Instrumentation%20\&body=Dear%20Illumina%20Technical%20Support,%0D%0A%0D%0A)*.* |
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