immersion fluid options for long timelapses with long working distance water objective

[Sylvie Le Guyader]

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Hi Pablo and everyone

I forward Steve Cody's answer to the list about this topic.
We have used his set up successfully on our Nikon Ti.

Med vänlig hälsning / Best regards

Sylvie

@@@@@@@@@@@@@@@@@@@@@@@@
Sylvie Le Guyader, PhD
Live Cell Imaging Facility Manager
Karolinska Institutet- Bionut Dpt
Hälsovägen 7C,
14157 Huddinge, Sweden
mobile: [phone]
LCI website
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-----Original Message-----
From: Confocal Microscopy List <[hidden email]> On Behalf Of Stephen Cody
Sent: 06 June 2019 07:21
To: [hidden email]
Subject: Re: Water dispenser pump settings

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Dear Darren,

I have used for many years a home made device. When grappling with this problem, I found it easier to just add excess water, ensure there was a large lip to catch excess water, and provide a scavenge pump to remove excess water. In this way you avoid trying to balance water evaporation rates and pump rates. You just pump more than enough. In the link you can download a pdf to show how you can make your own. I mostly now use a cut down GELoader tip to apply the water https://online-shop.eppendorf.com/OC-en/Manual-Liquid-Handling-44563/Pipette-Tips-44569/GELoader-PF-59967.html
,
and as a scavenge I use a bent 18 gauge "Drawing Up Needle" (eg.
https://www.signet.net.au/store/products/51914/terumo-neolus-needle-drawing-up-18-g-x-1-5-100-per-box
 )

If you make one of these devices and publish I'd appreciate it if you acknowledge "Stephen H. Cody, Monash Micro Imaging".
Thanks.

https://cloudstor.aarnet.edu.au/plus/index.php/s/thLe6ZNrA7C2FEz

All the best
Steve

On Sat, 11 Mar 2017 at 01:40, Bozzo Luigi <[hidden email]> wrote:

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>
> Dear Darren,
>
> I agree with Gabor and you that it is not straightforward to determine
> the amount of water to pump (we too have the Bartel water immersion
> dispenser for our SP8).
>
> In our case, for the evaporation we have found that with 250uL of
> water we can keep the focus for 3 hours in one position. Consider that
> our scope is inside a box of 37°C.
>
> For the accidental spilling water, Leica gave us a highly absorbent
> ring of foam (replaceable). We can wrap the foam around the water
> dispenser cap. Around 2cm of this ring of foam can absorb the entire
> volume of water contained in the dispenser (around 15ml).
>
> I take this opportunity to add a side comment. Recently, I saw a demo
> of the PerkinElmer Operetta (no commercial interest!). The system was
> equipped with a water immersion dispenser that I found simple and
> efficient. The water (coming from a 500ml bottle) was pumped out at
> the tip of the objective and the spilling water was collected by a
> collar and sent to another 500ml bottle.
>
> With this kind of dispenser no need to fine tune the water flux to
> match the loss due to evaporation, room temp, humidity, N of positions, etc..
> You can just pump a fixed volume for each position without worries for
> the water in excess.
>
> Best,
>
> Luigi
>
>
> On 09.03.2017 16:07, Darren Thomson wrote:
> > *****
> > To join, leave or search the confocal microscopy listserv, go to:
> > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> > Post images on http://www.imgur.com and include the link in your
> posting.
> > *****
> >
> > Thanks Gabor, it is indeed not straight-forward.
> >
> > Could you share your settings as a starting point for us?
> >
> > This message actually prompted a useful response from Leica:
> > As a starting point for experiments using the water dispenser I
> > would
> pump every half of an hour 1 sec with the settings 100V 100Hz. Using
> the MatrixScreener, where pump positions can be defined, I would pump
> roughly every third row of a multiwellplate with 96wells. For the
> movement of the stage I would set it to very slow values concerning speed and acceleration.
> This ensures, that the waterimmersion drop is following the stage movements.
> >
> > Cheers,
> >
> > Darren
> >
> > -----Original Message-----
> > From: Confocal Microscopy List
> > [mailto:[hidden email]]
> On Behalf Of Csúcs Gábor
> > Sent: 09 March 2017 14:34
> > To: [hidden email]
> > Subject: Re: Water dispenser pump settings
> >
> > *****
> > To join, leave or search the confocal microscopy listserv, go to:
> > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> > Post images on http://www.imgur.com and include the link in your
> posting.
> > *****
> >
> > Dear Darren,
> >
> > Not as defense of Leica, but our experience shows that it is indeed
> extremely difficult to give a detailed guideline how those values
> should be set. In fact it is not only the wettability of the plates
> and the temperature, but also the room /environment humidity, how many
> positions you are recording are all factors that count. Our solution
> for the was indeed "trial and error". On the other hand I agree with
> you (and share your frustration) that Leica didn't put much effort in
> documenting the properties etc of the system.
> >
> > Greetings    Gabor
> >
> > -----Original Message-----
> > From: Confocal Microscopy List
> > [mailto:[hidden email]]
> On Behalf Of Darren Thomson
> > Sent: Thursday, March 9, 2017 12:45 PM
> > To: [hidden email]
> > Subject: Water dispenser pump settings
> >
> > *****
> > To join, leave or search the confocal microscopy listserv, go to:
> > http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy
> > Post images on http://www.imgur.com and include the link in your
> posting.
> > *****
> >
> > Hello All,
> >
> > Is anyone using the Bartels Water Immersion Micro Dispenser on Leica
> water immersion lenses?
> > I'm looking for ball park settings to use on a 63x water immersion
> > lens
> on our SP8 confocal.
> > The manual has 1 setting, which causes spilling (9sec, 75 volts,
> > every 4
> mins @ 37D) at 1 position.
> >
> > Specifically, settings for 24/96 well plate visiting at 37D would be
> useful.
> >
> > I'm maybe over-complicating things by trying to match the dispensing
> rate with the evaporation rate. Then adding the drag of plate movement
> and intervals on top of that... Leica don't have any recommendations
> and ship the product without support "you just have to try it out at
> different settings" they say.
> >
> > Any suggestions would be most welcome!
> > Many thanks for your help,
> >
> > Dr Darren Thomson | Experimental Officer Manchester Fungal Infection
> Group (MFIG) University of Manchester
> > Tel: (+44) (0)161 306 2457
> > Email: [hidden email]
>
> --
> Luigi Bozzo, PhD
> Bioimaging and Optics Platform (PT-BIOP) Ecole Polytechnique Fédérale
> de Lausanne (EPFL) Faculty of Life Sciences Station 15, AI 0140
> CH-1015 Lausanne
>
> Phone: [phone]
> http://biop.epfl.ch/
>

--
Stephen H. Cody


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