☑️C12 Assembly and Sample Prep

How to put together your Omni-Stainer™ C12 system and prepare samples for staining

Step 1 - Create Humid Environment: add 30 mL deionized water to the Omni-Stainer™ C12 base

This helps to maintain a humid environment and prevents evaporation

Step 2 - Construct Flow Cell and Place in Base

You can create a flow cell in two methods:

1. [Shown in video below] Add ~100 ul of PBS on the Support Pad, and then place the coverslip (sample side down) onto the support pad. Adjust the coverslip ensuring it aligns with the lower boundary of the Support Pad. If there are air bubbles, sliding the coverslip such that bubble is released and then sliding it back after adding a bit more of the liquid allows to "expel" the bubbles

2. Align the coverslip and Support Pad "underwater" within a PBS pool, this technique that demands less skill to avoid air bubbles - however, the delicacy and clearness of coverslips occasionally make this less suitable for the C-style flow cell than for the S-style.

https://vimeo.com/751961194vimeo.com

A flow cell can be made with a drop of PBS (as in this video) or by constructing a flow cell within a reservoir of PBS

Preparing C type flow cell

Both the S (slide) and the C (coverslip) Omni-Stainers utilize "capillary liquid exchange " concept which is based on creating a thin layer of staining solution evenly covering the sample. The space where the sample is facing the exchangeable reactive staining solutions is called a "flow cell". To create the flow cell in C type Omni-Stainer the coverslip with the sample facing down needs to placed onto a Support Pad.

This can be achieved in a number of ways – the easiest one by doing it under the layer of PBS or any such compatible buffer inside a tip box lid or a beaker. As confidence and experience are built this can also be done by placing a drop of compatible buffer on top of the Support Pad and then covering the drop with a coverslip such that bubbles are avoided. In the case bubbles are encountered – sliding the coverslip such that bubble is released and then sliding it back after adding a bit more of the liquid allows to "expel" the bubbles

Preparing different sample-types

• FFPE sections: a sample needs to be baked, deparaffinized with Xylenes or a suitable alternative and re-hydrated. If HIER is needed, at this time we recommend it to be. It is possible to do HIER directly in Parhelia Omni-stainer using the Thermal Sheath, but that functionality is experimental and hasn’t been fully tested. Please reach out to us directly at [email protected] if you would like to try running HIER with Parhelia Omni-stainer.

• Fresh frozen sections: sample needs to be fixed and re-hydrated

• Cell culture/cell spreads: samples needs to be fixed and re-hydrated

Step 3 - Confirm Capillary Connection

It is important to verify that the coverslip aligns perfectly with the lower boundary of the Support Pad and makes contact with the wicking pillar. Confirm the existence of a capillary connection between the staining chamber and the wicking pillar, which essentially entails a liquid droplet linking the two components. If that’s not the case, dispense 100ul of PBS onto the contact point between the wicking pillar and the support tile (see diagram below).

Step 4 - Adhere moisture barrier paper to the lid's bottom, wet it with 10 mL of deionized water

Make sure the holes are aligned if using automation lid

Humidity barrier placement for manual lid

Step 5 - Put the lid on the base, matching slanted corners

Align cut diagonal corner of lid and base

Full Assembly

Insert sample, then put on lid (automation lid shown)

Complete Assembly (manual lid, 12 support pads)

C12 Automation Specs

C12 Manual Specs

Using a Thermal Sheath?

Thermal Sheath & Temperature-controlled incubations

Done! You're now ready to choose Manual or Automated mode and launch the demo!