Saturday, September 11, 2021

Scale up (or go to lab less!) with 384 well plates on your LC!

You probably already knew this, but until I didn't until recently. Every EasyNLC that I've ever used has been set up with 6x8 autosampler vials by the previous owner. I guess I've never had a new one, which... come to think of it... might have something to do with why I've been so mean about them, they're pretty finicky about being abused.... oh well... well, if you didn't know you can set them up with plates!  

Even if you use long gradients I bet there have been times where you've made tough decisions with 6x8 about either going in over the holidays or skipping some things because 48 x 3 hours still isn't all that long. We've been using 96 well plates, but with 30 minute gradients I've made hard decisions more than once. 

ENTER THE 384 well plate! 

As always don't take my word for any of this, where did you get this information? The internet? 

This is what I'm using

384 well plates (there are less expensive options, but there is some weird virus thing still disrupting stuff, some other 384 well plates that I've ordered for sample prep have been backordered for 5 weeks)

Webseal tape

Your EasyNLC probably already has an option for 16x24 plates. Woo! You're done! 

(I'm joking! Stop! You'll break your needle! I was going to put this below the picture but had a fit of conscience.) 

My A1 position was around B2.4. I don't know if I could have ran it without calibrating first, but if I had, my needle would have pushed right into the space between B2 and B3. 

You'll need this manual if you have an EasyNLC1200 (WARNING. DIRECT PDF LINK.)

You'll also want a flashlight (or a fancy phone with a flashlight. I made my Siri British somehow and I have to say "Siri, turn on the torch". In the basement here it says "working on it" then "working on it" then "something went wrong" in a snarky Alex Kingston need a flashlight.

As shown in the top picture, you need to remove the steel plate on the left side of your LC. The 3 screws are snap screws. Turn them counter clockwise 90 degrees and they'll pop free and the plate will come off. 

Install your 384 well plate (without tape on it, unsealed!) and start the calibrate procedure (the one in the middle of the screen)

The calibration isn't a complex one like an OpenTrons you just need to position 2 points. The bottom of your A1 well and the bottom of the well at the complete opposite corner of the plate.

If you touch the A1 position on the screen the needle will go somewhere near there, now you need to manually locate the bottom of the A1 well. 

You can see the A1 well perfectly so that one is easy. Use the extremely European touch screen controls (needle in/out is not into your well, it's into your instrument. you'll see what I mean) to navigate your needle to above the A1 and down until it touches. 

Now -- Take a picture of your needle coordinates. My A1 was this, I think, but what I care about is the depth of the needle in A1 because you can't see into the far opposite well (I don't know what that one is called yet, I only filled my first plate to N6) 

NOW you can capture your coordinate! Hit the green well above the opposite A1 and repeat the process. Get the flashlight. If you have bifocals, get those too! 

Without having the picture of the depth of A1 this will take forever because the plate will move just a little and you'll freak out that you hit bottom and will be afraid to go further. I verified that the bottom was just about perfectly the same on both sides by raising the needle 0.1 arbitrary units and finding just a little wiggle room in the plate). 

Now, I'll probably keep using this tape that I linked above, there is a very satisfying "POP" sound when the needle goes through it, but it does leave a permanent hole in the top, so you probably can't inject more than once out of a well unless you've filled a blank. Anything you worry about evaporating will probably evaporate, other than that, I'm up to L and the QA sample looks legit. Everything is working.

Oh crap. Almost forgot. 

You'll need to adjust your plate in your instrument configuration so it knows you did this. On a Thermo instrument it's the little double gear thing probably (which is now an App in Windows 10. I hope it makes this 1990s program feel totally hip or something).

On the Brukers it is one of the circled things. I always try them both first. 

You'll need to just let the queue know that you've got a 16x24 installed! Now you're done! 

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