# Third Party equipment frequently asked questions for the Illumina Protein Prep Automation System (IPPAS)

**What ancillary equipment is needed? (Note: quantities needed per 96 samples are listed in the Product Documentation.)**

Equipment (or equivalents thereof):

* Quad Microplate Shaker-Incubator Thermoshaker (VWR 97025-554)
* 8-channel screw cap decapper (Thermo 4105MAT)
* Thermal cycler (Cole-Parmer EW-93945-12, EW-93945-22, EW-93945-02, OR Bio-Rad 12015392)
* Axygen Silicone 96 Round Well Compression Flat Mat for Microplate (Fisher Scientific CM-FLAT)
* DynaMag-2 Magnet (Thermo 12321D)
* BioMicroLab Scan Barcode Reader (SPT LabTech, catalog # BMLScan)
* BioMicroLab Thaw rack thawing station (SPT LabTech, catalog # BML-RTS)
* BioShake XP - bench-top shaker (QInstruments, catalog # 1808-0505)
* Optional: Biotek Cytation 5 Cell Imaging and Multimode Reader (Agilent CYT5MFAV-SN)

The customer should consult the Product Documentation for a full list of required equipment and consumables.

**What are the vortex/shake and centrifuge settings during the automation protocol?**

**Script 1: Sample prep**

**PLASMADIL/SERUMDIL**: centrifuge plate 280 x g for 15 seconds

**Matrix Rack**: centrifuge at 1000 x g for 1 minute

**Binding**: shake for 3.5 h at 28C at 850 rpm

**Script 2: Proteomics Assay**

**Magnetic Beads**: shake until resuspended (minimum 60 seconds)

**MAGBEAD/MBPREP/MBBLOCK**: if splashing, centrifuge at 280 x g for 15 seconds

**Proteomics Probe Plate (PPP):** shake at 1350 rpm for 1 minute, centrifuge at 280 x g for 15 seconds

**Tag Reagent:** vortex for 5 seconds

**Hyb plate:** shake at 1350 rpm for 1 minute, centrifuge at 280 x g for 15 seconds

**Hybridization:**

**Hyb plate (if frozen):** centrifuge at 280 x g for 15 seconds, replace seal, shake at 1350 rpm for 1 minute. If splashing occurs, centrifuge at 280 x g for 15 seconds

**Script 3: Hyb Capture for NGS**

**Proteomics Amp Plate (PAP):** once thawed, shake at 1350 rpm for 1 minute, centrifuge at 280 x g for 15 seconds

**Indexes**: once thawed, shake at 1350 rpm for 1 minute, centrifuge at 280 x g for 15 seconds

**Proteomics Hyb Plate (PHP):** once thawed, centrifuge at 280 x g for 15 seconds, shake at 1800 rpm for 3 minutes

**Note:** Index PCR until the end of the protocol are performed manually. Follow protocol guidance for shake/vortex and centrifugation steps.

**How does the decapper work? What does it look like in action? Can the customer order more caps or another cap carrier?**

* The decapper looks similar to a multichannel pipette. It has square pegs that go into the square holes of the matrix sample tubes. It is strongly recommended to have one.
* 1 order of the decapper comes with 1 cap carrier. Labs can purchase additional cap carriers ([Fisher 11577429](https://www.fishersci.fi/shop/products/matrix-screwtop-tube-caps-cap-trays-cap-inserts/11577429), below), additional tubes, or caps if desired.

![Thermo Scientific Matrix ScrewTop Tube Caps, Cap Trays and Cap Inserts:Tubes,](https://761066130-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FGM9W2DuBTgEXv1ClCm8H%2Fuploads%2Fgit-blob-f3daaacb76e6262ff41cd69f3b444b878377371d%2Fimage1.jpg?alt=media)

**Is the shaker for SOMAmer binding step on-deck or off-deck? What does it look like?**

The shaker is the Quad Microplate Shaker-Incubator Thermoshaker and it is off-deck.

![](https://761066130-files.gitbook.io/~/files/v0/b/gitbook-x-prod.appspot.com/o/spaces%2FGM9W2DuBTgEXv1ClCm8H%2Fuploads%2Fgit-blob-181267c950fd1bb2c3bd9e9b07934f7b2a14cef1%2Fimage2.png?alt=media)

**Is a water bath required for reagents in 9.5k assay?**

Yes, it is required, but not required for all components of the assay.

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| *For any feedback or questions regarding this article (Illumina Knowledge Article #9799), contact Illumina Technical Support* [*techsupport@illumina.com*](mailto:techsupport@illumina.com?subject=Question%2FFeedback%20Regarding%20Illumina%20Knowledge%20Article%20#000009799%20-%20Library%20Preparation%20\&body=Dear%20Illumina%20Technical%20Support,%0D%0A%0D%0A)*.* |