TITLE: Automated transfer of inactivated virus samples from individual 2ml tubes to 96-well plate
AUTHORS: Efthymios Fidanis, Sophia Ward, Laura Cubitt, Amelia Edwards, Debbie Hughes, Michael Hubank, Jerome Nicod

[DESCRIPTION]
<div class = "text-blocks"><div class = "text-block"><span style = "font-weight:bold;">Purpose of examination / Clinical relevance</span></div><div class = "text-block"><div class = "justify" style = "text-align:left">At the end of 2019, several pneumonia cases were reported in Wuhan, China and the pathogen was confirmed as a new viral strain. World Health organization has named the newly identified coronavirus as 2019-nCoV, also known as SARS-Cov-2. The disease developed into a dangerous pandemic, posing major challenges to the NHS. Although more research is necessary to better understand the virus, in response to the emergency, simple and rapid testing is essential to identify the virus in infected individuals. This will aid the implementation of efficient interventions to contain the spread, and distinguish healthcare workers who have been infected, and are required to self-isolate, from those showing similar symptoms but which are not 2019-nCoV associated. The latter category may continue to work, alleviating stress on hard-pressed healthcare resources. 2019-nCoV is an RNA virus, and the diagnostic tests detect viral RNA in swabs from patient airways using a reverse transcriptase PCR assay. Samples are submitted to HSL, an accredited reporting laboratory, and transferred to the Crick for testing. The first step of the process is sample receipt at the Crick. This SOP describes the transfer of inactivated virus into plates for RNA extraction.</div></div><div class = "text-block"><div class = "justify" style = "text-align:left"></div></div><div class = "text-block"><div class = "justify" style = "text-align:left"><span style = "font-weight:bold;">Principles of Examination</span></div></div><div class = "text-block">The assay uses the Hamilton robot to transfer 150 μl of 93 samples of inactivated viral media samples from Barcoded 2 ml tubes (Cat no. 72.694.005) to Barcoded 96 well Nunc plates (Cat no. 260251) maintaining sample identity.</div><div class = "text-block">--</div><div class = "text-block">The authors wish to thank Heather Ringrose for support with the Hamilton liquid handlingworkstation.</div></div>

[STEPS]
?. [Transfer of inactivated virus to 96 well plates]
Sample tubes will arrive from the CL2/3 facility after virus inactivation at . Working in the reception area (in a separate room), wipe outside of tube with alcohol wipe, invert tube twice, and place in centrifuge. Pulse spin tubes.
0 Room temperature
?. [Transfer of inactivated virus to 96 well plates]
Remove tubes from centrifuge, place in new box (10x10) and transfer to automation lab.
?. [Transfer of inactivated virus to 96 well plates]
Unscrew 2 ml tubes containing sample and discard lid. Place open 2 ml tubes in Hamilton rack (Cat no. SMP_CAR_32_A00 with blue 2 ml tube inserts) with barcode facing forward as shown below:
The method has been installed on both our Hamilton STAR and STARlet instruments but this protocol only descibes the procedure on the STAR.
?. [Transfer of inactivated virus to 96 well plates]
Place rack in robot track position 45-47 (racks 1-3 respectively) depending on number of samples to process. Align the barcodes on the tubes to be visible in the inserts window. If processing 93 samples, leave rack 3, spaces 30, 31 and 32 empty for controls to be added later.
?. [Transfer of inactivated virus to 96 well plates]
Place an LPL barcode label on lower edge on the right-hand side of a 96-well Nunc (1.3 ml) plate (Cat no. 260251).
?. [Transfer of inactivated virus to 96 well plates]
Scan barcode into label printer to make an additional identical barcode for archiving box. Place in plate carrier (Cat no. PLT_CAR_L5MD_A00) in track position 49-54 on Hamilton robot.
?. [Transfer of inactivated virus to 96 well plates]
Access sample transfer protocol from ‘H’ button on Hamilton desktop.
?. [Transfer of inactivated virus to 96 well plates]
A pop-up window will give the option to use loading help. Always select ‘yes’.
?. [Transfer of inactivated virus to 96 well plates]
Follow guide using tip carrier with a tip rack in all 5 positions (can be empty, full or part used. Carrier cat no. TIP_CAR_480_AOO). Tip carrier placed in track positions 37-42.
?. [Transfer of inactivated virus to 96 well plates]
Check cover is closed as the final step, press OK, protocol will start and will transfer  to a specified position in the barcoded 96-well plate.
[inactivated virus solution]
?. [Transfer of inactivated virus to 96 well plates]
When program is complete, and the carriers are back in initial position, remove 96-well plate. Visually check volume in plate (this is easiest from below):
?. [Transfer of inactivated virus to 96 well plates]
Check .csv output for the transfer run for error messages. These should all be zero. If errors occur, report code to technical support. Upload .csv file into sample tracking app.
?. [Transfer of inactivated virus to 96 well plates]
Either: Cover plate and handling very carefully, pass plate immediately to RNA extraction team to be placed on BioMek Fx.Or: If plate is not to be used immediately, add plastic seal (cat no. AB-0558 located on bench next to Hamilton robot) and store 96-well Nunc plate at  in clear box labelled “to be extracted” (located on bench along from Hamilton robot) in ASF until extraction robot is available.
0 Room temperature
?. [Transfer of inactivated virus to 96 well plates]
Remove 2 ml tube containing residual sample from the Hamilton rack and cap with new coloured lid (located on bench in front of Hamilton and labelled as below).
?. [Archiving]
The app that used to perform the consolidation step will ask you to enter the barcode. Attach this barcode label to the box that will be used to archive the samples:
NOTE: This app is also used for archiving the RNA samples after extraction.
?. [Archiving]
When this step is carried out in Clarity LIMS, a database table is updated with the information. This step results in an entry appearing on the dashboard to let you know the samples are ready to be archived:
?. [Archiving]
An archiving app then displays the boxes and plates that are waiting to be archived, and gives you the chance to enter the storage information (see below):
?. [Archiving]
Once logged, the record disappears from this screen, and the archiving information will appear in the archive search app (see below):
?. [Archiving]
Place samples in freezer box labelled with barcode from  and store at .
-80 °C