Best practices for getting the lab back up and running after an extended shutdown

When returning to the lab after an extended shutdown or inactivity, it is important to make sure all lab resources are up and running and ready to be used. Below are best practices to check library prep, sequencing and microarray reagents and instruments to ensure a successful reopening.

Laboratory and Reagent Checks

Wipe down all lab surfaces with 10% bleach to remove any accumulated dust or contaminants.

• Pay particular attention to typical ‘hot spots’:

  • Benchtops used for processing amplified DNA

  • Door handles

  • Refrigerator and freezer door handles

  • Computer keyboard and mouse

  • Centrifuges, vortexers, and thermal cyclers

• Clean all equipment in any pre-amp areas (including pipettes)

• Refer to the bulletin Impact of ammonium-based cleaning products on sequencing run performance for additional details

Proper storage and handling of reagents, kits, and user-supplied materials should be followed according to documented procedures.

Check connectivity to network drives or servers used for file storage (output run folders, manifest files, cluster files, DMAPs) or data processing (IDAT repository).

Sequencing Instrumentation Considerations

Instrument System Guides: iSeq 100 | MiniSeq | MiSeq | NextSeq 500/550 | NextSeq 1000/2000 | HiSeq1000/1500/2000/2500 | HiSeq 3000/4000 | HiSeq X | NovaSeq 6000 | NovaSeq X Plus

For instruments that were left on:

1. Confirm the recommended maintenance/manual/standby washes were performed per the appropriate system guide.

2. Perform a full power cycle, following the steps in the system guide.

3. Leave system off for at least 5 minutes before powering back on.

Note: While the iSeq 100 and NextSeq 1000/2000 do not have onboard fluidics, a power cycle is recommended for these instruments as well.

For instruments that were turned off:

1. Power on the instrument and make sure that the control software initializes.

2. Perform two maintenance/manual washes to rehydrate the fluidics system (excluding iSeq 100 and NextSeq 1000/2000).

Note: If an additional maintenance/manual/standby wash using laboratory-grade water was not performed prior to instrument shutdown, perform three maintenance/manual washes to rehydrate the fluidics system, then run System Checks to test the fluidics delivery. System Check details are found in the appropriate system guides.

Library Preparation Considerations

Starting a New Library Prep

• When starting new library preparation, Illumina recommends reviewing the best practices for the specific library prep kit being used. This information is found on the appropriate product support page.

Proceeding to a Sequencing Run with Stored Libraries

• For any Illumina library that has been stored for longer than the recommended time, as specified in the reference guide, Illumina recommends repeating quality control (QC) and quantification prior to sequencing. Re-QC is beneficial because libraries can degrade during prolonged storage, leading to loss of overall yield. Recommended QC methods for most Illumina libraries are summarized in the Library quantification and quality control quick reference guide support bulletin.

For libraries requiring bead-based normalization, Illumina recommends storing the library at the last safe stopping point, just prior to the bead-based normalization step, as specified in the protocol guide. Libraries stored as double stranded DNA (dsDNA, before bead-based normalization) are more stable than libraries stored as single stranded DNA (ssDNA, after bead-based normalization). If a library has been stored long term after bead-based normalization, Illumina recommends repeating the bead-based normalization of the original dsDNA library.

Infinium Assay Considerations

• Refer to Infinium Assay Lab Setup and Best Practices and the appropriate Infinium Assay Reference Guide to make sure that all equipment and hardware are present and in good working condition.

• Note the duration and temperature recommendations for MSA plate storage stated in the ‘Safe Stopping Point’ portions of the Infinium Assay Reference Guide.

• Check stocks of user-supplied materials (eg, cap mats, 96-well plates, gloves, etc.) that may have been consumed by other groups using the space in the interim.

• Thoroughly wash glass back plates that may have accumulated dust or residue during the time of disuse. See the Technical Bulletin Improve Infinium data with these updated XStain glass back plate care recommendations for details.

Te-Flow and Water Circulator:

1. Confirm that there is no microbial growth in the water circulator and that the water is at the proper fill level.

   1. If microbial growth is present, follow the instructions in the Circulating Bath Manual (pg. 57 — 60) for cleaning the water circulator and refresh with algicide if needed.

   2. Do not use chlorine bleach to clean the water circulator.

2. Check the waste tubing lines of the Te-Flow and white reagent pan. Bleach can be squirted through the waste tubing lines leading to the waste carboy to clean them, if needed.

3. If the waste carboy has been emptied or moved, make sure it is back in its proper position, with an unimpeded, gravity-driven flow of waste from the Te-Flow Reagent pan to the waste carboy. Make sure that waste tubing is not kinked and there are no ‘uphill’ runs to the waste carboy.

Tecan/Illumina Automated Pipetting System (IAPS):

1. If the Tecan fluidics were stored dry, refill the system water carboy.

2. Empty the waste liquid container and disposable tip waste bag, if necessary.

3. Make sure that the Te-Flow is seated correctly in the pan, and that the Te-Flow is in the appropriate location on the Tecan deck.

4. Power on the Tecan.

5. Refer to the following sections in the Infinium Assay Lab Setup and Procedures Guide under Robot Usage and Maintenance:

   1. Preparing the Infinium Automated Pipetting System for Use (perform at least two bleach washes)

   2. Note: if the system was idle for a long time and/or dried out prior to shut down, multiple System Wash cycles are needed to purge all air from the fluidics.

   3. Testing Volume Accuracy of the IAPS Tips

iScan

1. Power on the scanner and make sure that the Archimedes drive appears on the PC before launching ICS software.

2. Clean the BeadChip carriers and iScan Nest (tray that carrier is placed on) with an Ethanol wipe to remove any dust or built up residue.

3. Turn on the scanner at least 30 minutes before use to give the lasers time to warm up.

For any feedback or questions regarding this article (Illumina Knowledge Article #3153), contact Illumina Technical Support [e-mail].