COVID Core Facility Training

[Jason M. Kirk]

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Hi Everyone,

As we continue to grapple with the long term effects of COVID-19 - those of us who operate shared instrumentation facilities have been finding ways to address the problem of how to provide the same high level of instruction while maintaining our commitment to social distancing and limited interpersonal contact.   While there is no one-size-fits-all solution – I wanted to share the route we have chosen in the hopes that it will provide some benefit to others.

Confocal microscopes are the bread and butter of our core – and we have created video-based training that so far has been doing a consistent job replicating our traditional confocal training.  The key for us has been creating high quality video content that replicates every aspect of our in-person training session.  The main issue I’ve noticed with much of the video based training publicly available is that it is nowhere near as in-depth as we traditionally get during our one-on-one sessions.  Creating detailed content that is specific to our instruments has allowed us to almost completely remove ourselves from the initial training.

We’ve been requiring users to watch an introductory video as well as the full training video before arriving for their ‘in-person’ events.  When they arrive we give them a quick overview of the core and then have them go through the full resolution video on a monitor right next to the instrument.  This way they can follow along and start/stop the instruction to move at their pace.  A second session is then used to gauge how well the user is absorbing the material.

It has been quite well received – and I’ve posted the full 4K videos for our first instrument (Zeiss LSM 880 with Airyscan) to Youtube here:

https://www.youtube.com/channel/UCVOV2V_G20MvT79gfXBYO5A

We will be adding videos as we expand training for our other instruments – as well as more advanced techniques as our users request them.

Of course these videos are specific to our instruments – but I hope they can be useful to others looking to do similar things.

Cheers!

-Jason

-------------------------------------------------------------------
Jason M. Kirk
Technical Director, Optical Imaging & Vital Microscopy Core (OiVM)
Baylor College of Medicine
Ph: 713.798.6486
Email: [hidden email]
http://www.bcm.edu/oivm

[Ariel, Pablo]

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You've made some very high quality videos; thanks for sharing them with the community.

In our core, we've been doing something very similar, and have also had good results. We do a few things slightly differently:
1) We ask users to watch the videos in advance, not during the training. Like yours, those videos are also very detailed and specific to our instruments.
2) For our more complicated systems (laser scanning confocals, Lavision light-sheet), we provide detailed written instructions that they can use to follow along during the video, as well as bring to the training.
3) We have an online quiz they have to complete before the in-house training to make sure they watched the videos and got the most important points.
4) We check that their sample has some sort of reasonable signal before the in-house training.
5) The in-house training involves a brief interaction reinforcing the steps for setting up their sample. The rest is remote, via zoom, with screen sharing and remote control. The "remote" staff member is an office nearby and can walk over and help if they get stuck and the issue is not resolvable via zoon.

This training process has worked very well. With few exceptions, people come much better prepared and the training is smoother. I will probably keep the requirement to watch the training videos when this mess is over.

In case anyone finds anything useful there, here are our video playlists:
https://www.youtube.com/channel/UCTqLyJ-2uBIl0hHV9mD4pCQ/playlists
The production quality is significantly worse than what Jason's core has developed, but I think some of the playlists may be useful to others. In addition to instrument trainings, we have some general lectures and software tutorials. I also have a series of maintenance videos for a LaVision Ultra-II light-sheet, mainly as a reference for my own staff.

Best,
Pablo


Pablo Ariel, Ph.D.
Assistant Professor
Director of the Microscopy Services Laboratory
Brinkhous-Bullitt Bldg B04
Department of Pathology and Laboratory Medicine
University of North Carolina at Chapel Hill
http://www.med.unc.edu/microscopy
Tel: [phone]




-----Original Message-----
From: Confocal Microscopy List <[hidden email]> On Behalf Of Jason M. Kirk
Sent: Monday, December 7, 2020 2:14 PM
To: [hidden email]
Subject: COVID Core Facility Training

*****
To join, leave or search the confocal microscopy listserv, go to:
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*****

Hi Everyone,

As we continue to grapple with the long term effects of COVID-19 - those of us who operate shared instrumentation facilities have been finding ways to address the problem of how to provide the same high level of instruction while maintaining our commitment to social distancing and limited interpersonal contact.   While there is no one-size-fits-all solution – I wanted to share the route we have chosen in the hopes that it will provide some benefit to others.

Confocal microscopes are the bread and butter of our core – and we have created video-based training that so far has been doing a consistent job replicating our traditional confocal training.  The key for us has been creating high quality video content that replicates every aspect of our in-person training session.  The main issue I’ve noticed with much of the video based training publicly available is that it is nowhere near as in-depth as we traditionally get during our one-on-one sessions.  Creating detailed content that is specific to our instruments has allowed us to almost completely remove ourselves from the initial training.

We’ve been requiring users to watch an introductory video as well as the full training video before arriving for their ‘in-person’ events.  When they arrive we give them a quick overview of the core and then have them go through the full resolution video on a monitor right next to the instrument.  This way they can follow along and start/stop the instruction to move at their pace.  A second session is then used to gauge how well the user is absorbing the material.

It has been quite well received – and I’ve posted the full 4K videos for our first instrument (Zeiss LSM 880 with Airyscan) to Youtube here:

https://www.youtube.com/channel/UCVOV2V_G20MvT79gfXBYO5A

We will be adding videos as we expand training for our other instruments – as well as more advanced techniques as our users request them.

Of course these videos are specific to our instruments – but I hope they can be useful to others looking to do similar things.

Cheers!

-Jason

-------------------------------------------------------------------
Jason M. Kirk
Technical Director, Optical Imaging & Vital Microscopy Core (OiVM) Baylor College of Medicine
Ph: 713.798.6486
Email: [hidden email]
http://www.bcm.edu/oivm

[samuel connell]

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Please pardon the brief commercial interruption:

I wanted to share with this community that 3i has developed an adaptive
optics module. We are presenting a tech talk on this new development on
Wednesday at Cell Bio 2020. Follow the link below if you are attending this
meeting and would like to register. If you'd like to learn more outside of
this setting, let me know and we can follow up.

C-Shaper is a turnkey hardware and software solution to bring easy-to-use
adaptive optics to the Marianas live cell imaging platform. Paired with
Yokogawa CSU-W1 or CSU-X1, C-Shaper enables aberration-corrected imaging
across multiple objectives - restoring objective performance, boosting
signal-to-noise, and increasing imaging depth.

C-Shaper: A New Approach for Adaptive Optics in Microscopy
<https://cell-bio-virtual2020.pathable.co/meetings/virtual/QdGKQYFD5DoSEFtFa>
Time: 10:00 AM - 10:45 AM EST on Wednesday, December 9
Speaker: Omer Tzang, Ph.D., Research and Development Scientist

https://go.intelligent-imaging.com/c-shaper-adaptive-optics

Cheers,
-
Sam

Samuel Connell
Director of Sales

Intelligent Imaging Innovations (3i)
3575 Ringsby Ct, Suite 102
Denver, CO 80216 USA
1-720-437-6926
www.intelligent-imaging.com
Twitter: @samuelconnell

[G. Esteban Fernandez]

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Just looked at the Intro. video and it is superb!  Touches on everything I
tell people during training, with really great illustrations to drive home
the points.  Thank you Jason.

-Esteban

On Mon, Dec 7, 2020 at 11:14 AM Jason M. Kirk <[hidden email]> 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.
> *****
>
> Hi Everyone,
>
> As we continue to grapple with the long term effects of COVID-19 - those
> of us who operate shared instrumentation facilities have been finding ways
> to address the problem of how to provide the same high level of instruction
> while maintaining our commitment to social distancing and limited
> interpersonal contact.   While there is no one-size-fits-all solution – I
> wanted to share the route we have chosen in the hopes that it will provide
> some benefit to others.
>
> Confocal microscopes are the bread and butter of our core – and we have
> created video-based training that so far has been doing a consistent job
> replicating our traditional confocal training.  The key for us has been
> creating high quality video content that replicates every aspect of our
> in-person training session.  The main issue I’ve noticed with much of the
> video based training publicly available is that it is nowhere near as
> in-depth as we traditionally get during our one-on-one sessions.  Creating
> detailed content that is specific to our instruments has allowed us to
> almost completely remove ourselves from the initial training.
>
> We’ve been requiring users to watch an introductory video as well as the
> full training video before arriving for their ‘in-person’ events.  When
> they arrive we give them a quick overview of the core and then have them go
> through the full resolution video on a monitor right next to the
> instrument.  This way they can follow along and start/stop the instruction
> to move at their pace.  A second session is then used to gauge how well the
> user is absorbing the material.
>
> It has been quite well received – and I’ve posted the full 4K videos for
> our first instrument (Zeiss LSM 880 with Airyscan) to Youtube here:
>
> https://www.youtube.com/channel/UCVOV2V_G20MvT79gfXBYO5A
>
> We will be adding videos as we expand training for our other instruments –
> as well as more advanced techniques as our users request them.
>
> Of course these videos are specific to our instruments – but I hope they
> can be useful to others looking to do similar things.
>
> Cheers!
>
> -Jason
>
> -------------------------------------------------------------------
> Jason M. Kirk
> Technical Director, Optical Imaging & Vital Microscopy Core (OiVM)
> Baylor College of Medicine
> Ph: 713.798.6486
> Email: [hidden email]
> http://www.bcm.edu/oivm
>

[Arvydas Matiukas]

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*****

Hi Jason,

I second Esteban's congrats with an excellent confocal training video that covers all major areas that a new user should be familiar with. It is especially useful for the people using Zeiss confocals. I only wonder why you did not show "Smart setup" function and Fluorophore emission spectra Database. In my experience reusing previous experiment/image configurations and Smart setup are the two main  features that allow obtaining better and faster images for the beginner users.

Best regards,
Arvydas
*****************************



Arvydas Matiukas, Ph.D.

Director of  Neuroscience  Microscopy Core

Manager of NRB Shared Research Equipment

SUNY Upstate Medical University


Email: [hidden email]

________________________________
From: Confocal Microscopy List <[hidden email]> on behalf of G. Esteban Fernandez <[hidden email]>
Sent: Monday, December 7, 2020 3:45 PM
To: [hidden email] <[hidden email]>
Subject: [EXTERNAL] Re: COVID Core Facility Training

*****
To join, leave or search the confocal microscopy listserv, go to:
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Post images on https://urldefense.com/v3/__http://www.imgur.com__;!!GobTDDpD7A!eGt5QMSRfbyPXhScsXVP_Vt1SKw-wlSzDqa164xVomRbxTnkoxJNw539nN11tS5RhA$  and include the link in your posting.
*****

Just looked at the Intro. video and it is superb!  Touches on everything I
tell people during training, with really great illustrations to drive home
the points.  Thank you Jason.

-Esteban

On Mon, Dec 7, 2020 at 11:14 AM Jason M. Kirk <[hidden email]> wrote:

> *****
> To join, leave or search the confocal microscopy listserv, go to:
> https://urldefense.com/v3/__http://lists.umn.edu/cgi-bin/wa?A0=confocalmicroscopy__;!!GobTDDpD7A!eGt5QMSRfbyPXhScsXVP_Vt1SKw-wlSzDqa164xVomRbxTnkoxJNw539nN2sMI3TfQ$
> Post images on https://urldefense.com/v3/__http://www.imgur.com__;!!GobTDDpD7A!eGt5QMSRfbyPXhScsXVP_Vt1SKw-wlSzDqa164xVomRbxTnkoxJNw539nN11tS5RhA$  and include the link in your posting.
> *****
>
> Hi Everyone,
>
> As we continue to grapple with the long term effects of COVID-19 - those
> of us who operate shared instrumentation facilities have been finding ways
> to address the problem of how to provide the same high level of instruction
> while maintaining our commitment to social distancing and limited
> interpersonal contact.   While there is no one-size-fits-all solution – I
> wanted to share the route we have chosen in the hopes that it will provide
> some benefit to others.
>
> Confocal microscopes are the bread and butter of our core – and we have
> created video-based training that so far has been doing a consistent job
> replicating our traditional confocal training.  The key for us has been
> creating high quality video content that replicates every aspect of our
> in-person training session.  The main issue I’ve noticed with much of the
> video based training publicly available is that it is nowhere near as
> in-depth as we traditionally get during our one-on-one sessions.  Creating
> detailed content that is specific to our instruments has allowed us to
> almost completely remove ourselves from the initial training.
>
> We’ve been requiring users to watch an introductory video as well as the
> full training video before arriving for their ‘in-person’ events.  When
> they arrive we give them a quick overview of the core and then have them go
> through the full resolution video on a monitor right next to the
> instrument.  This way they can follow along and start/stop the instruction
> to move at their pace.  A second session is then used to gauge how well the
> user is absorbing the material.
>
> It has been quite well received – and I’ve posted the full 4K videos for
> our first instrument (Zeiss LSM 880 with Airyscan) to Youtube here:
>
> https://urldefense.com/v3/__https://www.youtube.com/channel/UCVOV2V_G20MvT79gfXBYO5A__;!!GobTDDpD7A!eGt5QMSRfbyPXhScsXVP_Vt1SKw-wlSzDqa164xVomRbxTnkoxJNw539nN0cS1sfvw$
>
> We will be adding videos as we expand training for our other instruments –
> as well as more advanced techniques as our users request them.
>
> Of course these videos are specific to our instruments – but I hope they
> can be useful to others looking to do similar things.
>
> Cheers!
>
> -Jason
>
> -------------------------------------------------------------------
> Jason M. Kirk
> Technical Director, Optical Imaging & Vital Microscopy Core (OiVM)
> Baylor College of Medicine
> Ph: 713.798.6486
> Email: [hidden email]
> https://urldefense.com/v3/__http://www.bcm.edu/oivm__;!!GobTDDpD7A!eGt5QMSRfbyPXhScsXVP_Vt1SKw-wlSzDqa164xVomRbxTnkoxJNw539nN3DElmgDQ$
>

[Jason M. Kirk]

*****
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*****

Hi Arvydas,

I intentionally ignore the Smart Setup in ZEN for a few reasons.  The main reason is that it does a horrible job setting up sequential acquisitions.  It forces the design of the beampath to use frame-wise track switching – and then proceeds to design light paths that switch beam combiners and filters between tracks when it is completely unnecessary to do so.  The Frame mode imposes a ~500ms delay between tracks to allow for mechanical changes – which doesn’t sound like a lot but when you account for the delay between each channel plus Z, tile or whatever other multi-dimensional acquisition parameters the user requires it quickly adds up.  Over the course of an average 3D acquisition – manually refining your light path can reduce the total acquisition time by up to 50% vs. Smart Setup.

Smart Setup also programs extremely wide emission bands for each fluorophore which if left unmodified will most certainly lead to emission bleed.  

Another reason is that it completely ignores the airyscan detector as a viable 4th confocal detector.  So on our setup – we can do 4 color line switching between tracks for sequential imaging.
 
While I do agree that the beampath configuration has a steep learning curve for new users – I feel that glossing over this aspect of the confocal use would be doing them a disservice in the long run.  I don’t expect my users to manually design beampaths every time they use the instrument – once they’ve worked out their ideal parameters the Reuse and experiment configuration functionality is invaluable – but while they are learning I feel that it is important that they understand the level of control they have over the design of their acquisition.

-Jason

-------------------------------------------------------------------
Jason M. Kirk
Technical Director, Optical Imaging & Vital Microscopy Core (OiVM)
Baylor College of Medicine
Ph: 713.798.6486
Email: [hidden email]
http://www.bcm.edu/oivm

[Alison J. North-2]

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*****

Hi Jason,

Just want to say that my staff and I are 100% behind you on this one!  We have many users who do a ton of tiled, multicolour z-stacks, and the overall difference in time between the "Smart" Setup track and one that we have set up manually is unbelievable.  Not to mention the fact that, if I remember correctly, when you use Smart Setup to establish a single colour track for GFP on one of our systems, it decides to avoid using the GaAsP detector, for some bizarre reason.

And I also agree entirely that education about what is going on in the track should be a critically important part of training our users.  Many of them go off to buy their own confocal systems later, and it is really important that they learn how to get the best out of the system for themselves.

Thank you for making and sharing such great videos, I am hearing very positive comments about them from all over!
All the best,
Alison

________________________________
From: Confocal Microscopy List <[hidden email]> on behalf of Jason M. Kirk <[hidden email]>
Sent: Tuesday, December 8, 2020 10:26 AM
To: [hidden email] <[hidden email]>
Subject: Re: [EXTERNAL] Re: COVID Core Facility Training

*****
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*****

Hi Arvydas,

I intentionally ignore the Smart Setup in ZEN for a few reasons.  The main reason is that it does a horrible job setting up sequential acquisitions.  It forces the design of the beampath to use frame-wise track switching – and then proceeds to design light paths that switch beam combiners and filters between tracks when it is completely unnecessary to do so.  The Frame mode imposes a ~500ms delay between tracks to allow for mechanical changes – which doesn’t sound like a lot but when you account for the delay between each channel plus Z, tile or whatever other multi-dimensional acquisition parameters the user requires it quickly adds up.  Over the course of an average 3D acquisition – manually refining your light path can reduce the total acquisition time by up to 50% vs. Smart Setup.

Smart Setup also programs extremely wide emission bands for each fluorophore which if left unmodified will most certainly lead to emission bleed.

Another reason is that it completely ignores the airyscan detector as a viable 4th confocal detector.  So on our setup – we can do 4 color line switching between tracks for sequential imaging.

While I do agree that the beampath configuration has a steep learning curve for new users – I feel that glossing over this aspect of the confocal use would be doing them a disservice in the long run.  I don’t expect my users to manually design beampaths every time they use the instrument – once they’ve worked out their ideal parameters the Reuse and experiment configuration functionality is invaluable – but while they are learning I feel that it is important that they understand the level of control they have over the design of their acquisition.

-Jason

-------------------------------------------------------------------
Jason M. Kirk
Technical Director, Optical Imaging & Vital Microscopy Core (OiVM)
Baylor College of Medicine
Ph: 713.798.6486
Email: [hidden email]
https://urldefense.proofpoint.com/v2/url?u=http-3A__www.bcm.edu_oivm&d=DwIFaQ&c=JeTkUgVztGMmhKYjxsy2rfoWYibK1YmxXez1G3oNStg&r=RBx0-WJrAO5vwSOLNmFbqYvikvIZS5ns3-USwvMOuLo&m=csLzCzmvqRHbD7-N3787XLe_P7KtLGKiF07rZUseSn4&s=yiG4VdP0PBl_sE4ZBQlpmZNrIsLk58VV_demyQ2tnwg&e=

[Moulding, Dale]

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*****

Hi Jason,

These videos are fantastic. Thank you!  The intro will now be essential viewing for all my new users. I'll encourage all users (especially experienced ones) to watch them all as they should reverse any bad habits they've fallen in to. Inhope that now they've got to grips with using the system they should be able to absorb the info about how it actually works.

To smart set up or not? I teach new users to use smart set up, then show them that switching to line (for using live) and frame fast (for acquisition) is much much faster, but requires you to change a few settings. In the end I find that easier than setting all up from scratch. The user gets a helping hand from smart set up, then makes it smarter with a little fine tuning.

I think on the 880 software, smart set up offers a best  compromise that always switches by line?

Single and two channel smart setup seems to always put the lower wavelength to Ch1, and the second wavelength to Ch2. So never uses the GaAsP for a single channel image. I always find that strange, so again let the users decide if smart setup was smart enough.

Cheers

Dale

Dale Moulding PhD FRMS
ICH Light Microscopy Facility
Room W2.06, BDRC office
UCL Great Ormond Street Institute of Child Health
30 Guilford St
London WC1N 1EH
Mob: [phone]
Tel: [phone]
http://www.ucl.ac.uk/ich/core-scientific-facilities-centres/confocal-microscopy

________________________________
From: Confocal Microscopy List <[hidden email]> on behalf of Alison J. North <[hidden email]>
Sent: Tuesday, December 8, 2020 3:36:25 PM
To: [hidden email] <[hidden email]>
Subject: Re: [EXTERNAL] Re: COVID Core Facility Training

*****
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*****

Hi Jason,

Just want to say that my staff and I are 100% behind you on this one!  We have many users who do a ton of tiled, multicolour z-stacks, and the overall difference in time between the "Smart" Setup track and one that we have set up manually is unbelievable.  Not to mention the fact that, if I remember correctly, when you use Smart Setup to establish a single colour track for GFP on one of our systems, it decides to avoid using the GaAsP detector, for some bizarre reason.

And I also agree entirely that education about what is going on in the track should be a critically important part of training our users.  Many of them go off to buy their own confocal systems later, and it is really important that they learn how to get the best out of the system for themselves.

Thank you for making and sharing such great videos, I am hearing very positive comments about them from all over!
All the best,
Alison

________________________________
From: Confocal Microscopy List <[hidden email]> on behalf of Jason M. Kirk <[hidden email]>
Sent: Tuesday, December 8, 2020 10:26 AM
To: [hidden email] <[hidden email]>
Subject: Re: [EXTERNAL] Re: COVID Core Facility Training

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Hi Arvydas,

I intentionally ignore the Smart Setup in ZEN for a few reasons.  The main reason is that it does a horrible job setting up sequential acquisitions.  It forces the design of the beampath to use frame-wise track switching – and then proceeds to design light paths that switch beam combiners and filters between tracks when it is completely unnecessary to do so.  The Frame mode imposes a ~500ms delay between tracks to allow for mechanical changes – which doesn’t sound like a lot but when you account for the delay between each channel plus Z, tile or whatever other multi-dimensional acquisition parameters the user requires it quickly adds up.  Over the course of an average 3D acquisition – manually refining your light path can reduce the total acquisition time by up to 50% vs. Smart Setup.

Smart Setup also programs extremely wide emission bands for each fluorophore which if left unmodified will most certainly lead to emission bleed.

Another reason is that it completely ignores the airyscan detector as a viable 4th confocal detector.  So on our setup – we can do 4 color line switching between tracks for sequential imaging.

While I do agree that the beampath configuration has a steep learning curve for new users – I feel that glossing over this aspect of the confocal use would be doing them a disservice in the long run.  I don’t expect my users to manually design beampaths every time they use the instrument – once they’ve worked out their ideal parameters the Reuse and experiment configuration functionality is invaluable – but while they are learning I feel that it is important that they understand the level of control they have over the design of their acquisition.

-Jason

-------------------------------------------------------------------
Jason M. Kirk
Technical Director, Optical Imaging & Vital Microscopy Core (OiVM)
Baylor College of Medicine
Ph: 713.798.6486
Email: [hidden email]
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[Armstrong, Brian]

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Hi Jason and list, I would like to whole heartedly agree with Jason's advice below.

Cheers!

Brian Armstrong PhD
Associate Research Professor
Developmental and Stem Cell Biology
Diabetes and Metabolic Diseases
Director, Light Microscopy Core
Beckman Research Institute, City of Hope


-----Original Message-----
From: Confocal Microscopy List [mailto:[hidden email]] On Behalf Of Jason M. Kirk
Sent: Tuesday, December 08, 2020 7:26 AM
To: [hidden email]
Subject: Re: [EXTERNAL] Re: COVID Core Facility Training

[Attention: This email came from an external source. Do not open attachments or click on links from unknown senders or unexpected emails.]

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Hi Arvydas,

I intentionally ignore the Smart Setup in ZEN for a few reasons.  The main reason is that it does a horrible job setting up sequential acquisitions.  It forces the design of the beampath to use frame-wise track switching – and then proceeds to design light paths that switch beam combiners and filters between tracks when it is completely unnecessary to do so.  The Frame mode imposes a ~500ms delay between tracks to allow for mechanical changes – which doesn’t sound like a lot but when you account for the delay between each channel plus Z, tile or whatever other multi-dimensional acquisition parameters the user requires it quickly adds up.  Over the course of an average 3D acquisition – manually refining your light path can reduce the total acquisition time by up to 50% vs. Smart Setup.

Smart Setup also programs extremely wide emission bands for each fluorophore which if left unmodified will most certainly lead to emission bleed.  

Another reason is that it completely ignores the airyscan detector as a viable 4th confocal detector.  So on our setup – we can do 4 color line switching between tracks for sequential imaging.
 
While I do agree that the beampath configuration has a steep learning curve for new users – I feel that glossing over this aspect of the confocal use would be doing them a disservice in the long run.  I don’t expect my users to manually design beampaths every time they use the instrument – once they’ve worked out their ideal parameters the Reuse and experiment configuration functionality is invaluable – but while they are learning I feel that it is important that they understand the level of control they have over the design of their acquisition.

-Jason

-------------------------------------------------------------------
Jason M. Kirk
Technical Director, Optical Imaging & Vital Microscopy Core (OiVM) Baylor College of Medicine
Ph: 713.798.6486
Email: [hidden email]
https://urldefense.com/v3/__http://www.bcm.edu/oivm__;!!Fou38LsQmgU!7RA12lCPMIeC0hiP1ShVj29HW-Y_2J0uqZH74DoCoNzGH_L0sjNsbmbVVi8loZI$ 

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[Darran Clements]

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Hi Jason
Really good points - the Smart setup can take you to some fairly off-piste setups. However, I always train people using it then use the setup it produces as a learning experience for the new user to go through the reasons for why the smart setup is wrong, it usually lightens the mood and lets me delve into the finer points of how to use the microscope more effectively... filter specs, detector sensitivity, beam path descriptions etc. And now that LAS X has something very similar, it's not going away and it can be a useful prop for a really in depth training session... certainly a lot more illuminating than taking someone through a training session and their take-home message being "I can always use Apply/Reuse" for future imaging sessions, which is, unfortunately, all to common when you inherit users who haven't been adequately trained.
D

Wellcome-MRC  Stem Cell Institute
Jeffrey Cheah Biomedical Centre
Cambridge Biomedical Campus
University of Cambridge
Puddicombe Way
Cambridge
CB2 0AW






-----Original Message-----
From: Confocal Microscopy List <[hidden email]> On Behalf Of Jason M. Kirk
Sent: 08 December 2020 15:26
To: [hidden email]
Subject: Re: [EXTERNAL] Re: COVID Core Facility Training

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Hi Arvydas,

I intentionally ignore the Smart Setup in ZEN for a few reasons.  The main reason is that it does a horrible job setting up sequential acquisitions.  It forces the design of the beampath to use frame-wise track switching – and then proceeds to design light paths that switch beam combiners and filters between tracks when it is completely unnecessary to do so.  The Frame mode imposes a ~500ms delay between tracks to allow for mechanical changes – which doesn’t sound like a lot but when you account for the delay between each channel plus Z, tile or whatever other multi-dimensional acquisition parameters the user requires it quickly adds up.  Over the course of an average 3D acquisition – manually refining your light path can reduce the total acquisition time by up to 50% vs. Smart Setup.

Smart Setup also programs extremely wide emission bands for each fluorophore which if left unmodified will most certainly lead to emission bleed.  

Another reason is that it completely ignores the airyscan detector as a viable 4th confocal detector.  So on our setup – we can do 4 color line switching between tracks for sequential imaging.
 
While I do agree that the beampath configuration has a steep learning curve for new users – I feel that glossing over this aspect of the confocal use would be doing them a disservice in the long run.  I don’t expect my users to manually design beampaths every time they use the instrument – once they’ve worked out their ideal parameters the Reuse and experiment configuration functionality is invaluable – but while they are learning I feel that it is important that they understand the level of control they have over the design of their acquisition.

-Jason

-------------------------------------------------------------------
Jason M. Kirk
Technical Director, Optical Imaging & Vital Microscopy Core (OiVM) Baylor College of Medicine
Ph: 713.798.6486
Email: [hidden email]
http://www.bcm.edu/oivm

[Arvydas Matiukas]

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Hi Confocal Trainers,

    I think the  discussion and sharing training experience is getting really  interesting and useful.  I only would add not to confuse the training (and related video/materials) which is a process (from basic to advanced confocal) with setting up and implementing optimized imaging experiment that is the ultimate goal. Confocal software and its tools evolved a lot since 2000, and if it is not yet at the level of the self-driving car, it is getting quite user friendly and flexible (and even has different levels of menus/tools). Therefore I think it is useful at least to mention all available setup options/tools (along with their pros/cons) and let the user to choose the one that works best for her/him. I always assure trainees that any workflow is acceptable (and likely close to optimal) as long as it provides required (quality) images and measurements. Similarly for different math tasks: sometimes calculator, sometimes Excel is preferred (but for major discovery one may need a supercomputer!)

   Jason, Smart setup works better on our 780 machine because it does not have Airyscan. In my experience it provides  close to optimal optical setup for up to 2 (sometimes even 3) fluorophores with well separated Ex/Em spectra, and can be used as a quick template for further optimization. I agree that the Smart setup is not efficient with 4 dyes, multicolor z-stacks and other advanced experiments.
    You mentioned reducing signal bleed-though by narrowing emission bandwidth. I wonder how you teach to implement it: by using external spectral viewers/calculators, by comparing signals under different settings, by using the built-in spectra database (BTW, Smart setup estimates % of the bleed-through, etc).

Best,
Arvydas

________________________________
From: Confocal Microscopy List <[hidden email]> on behalf of Moulding, Dale <[hidden email]>
Sent: Tuesday, December 8, 2020 11:08 AM
To: [hidden email] <[hidden email]>
Subject: Re: [EXTERNAL] Re: COVID Core Facility Training

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Hi Jason,

These videos are fantastic. Thank you!  The intro will now be essential viewing for all my new users. I'll encourage all users (especially experienced ones) to watch them all as they should reverse any bad habits they've fallen in to. Inhope that now they've got to grips with using the system they should be able to absorb the info about how it actually works.

To smart set up or not? I teach new users to use smart set up, then show them that switching to line (for using live) and frame fast (for acquisition) is much much faster, but requires you to change a few settings. In the end I find that easier than setting all up from scratch. The user gets a helping hand from smart set up, then makes it smarter with a little fine tuning.

I think on the 880 software, smart set up offers a best  compromise that always switches by line?

Single and two channel smart setup seems to always put the lower wavelength to Ch1, and the second wavelength to Ch2. So never uses the GaAsP for a single channel image. I always find that strange, so again let the users decide if smart setup was smart enough.

Cheers

Dale

Dale Moulding PhD FRMS
ICH Light Microscopy Facility
Room W2.06, BDRC office
UCL Great Ormond Street Institute of Child Health
30 Guilford St
London WC1N 1EH
Mob: [phone]
Tel: [phone]
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________________________________
From: Confocal Microscopy List <[hidden email]> on behalf of Alison J. North <[hidden email]>
Sent: Tuesday, December 8, 2020 3:36:25 PM
To: [hidden email] <[hidden email]>
Subject: Re: [EXTERNAL] Re: COVID Core Facility Training

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Hi Jason,

Just want to say that my staff and I are 100% behind you on this one!  We have many users who do a ton of tiled, multicolour z-stacks, and the overall difference in time between the "Smart" Setup track and one that we have set up manually is unbelievable.  Not to mention the fact that, if I remember correctly, when you use Smart Setup to establish a single colour track for GFP on one of our systems, it decides to avoid using the GaAsP detector, for some bizarre reason.

And I also agree entirely that education about what is going on in the track should be a critically important part of training our users.  Many of them go off to buy their own confocal systems later, and it is really important that they learn how to get the best out of the system for themselves.

Thank you for making and sharing such great videos, I am hearing very positive comments about them from all over!
All the best,
Alison

________________________________
From: Confocal Microscopy List <[hidden email]> on behalf of Jason M. Kirk <[hidden email]>
Sent: Tuesday, December 8, 2020 10:26 AM
To: [hidden email] <[hidden email]>
Subject: Re: [EXTERNAL] Re: COVID Core Facility Training

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*****

Hi Arvydas,

I intentionally ignore the Smart Setup in ZEN for a few reasons.  The main reason is that it does a horrible job setting up sequential acquisitions.  It forces the design of the beampath to use frame-wise track switching – and then proceeds to design light paths that switch beam combiners and filters between tracks when it is completely unnecessary to do so.  The Frame mode imposes a ~500ms delay between tracks to allow for mechanical changes – which doesn’t sound like a lot but when you account for the delay between each channel plus Z, tile or whatever other multi-dimensional acquisition parameters the user requires it quickly adds up.  Over the course of an average 3D acquisition – manually refining your light path can reduce the total acquisition time by up to 50% vs. Smart Setup.

Smart Setup also programs extremely wide emission bands for each fluorophore which if left unmodified will most certainly lead to emission bleed.

Another reason is that it completely ignores the airyscan detector as a viable 4th confocal detector.  So on our setup – we can do 4 color line switching between tracks for sequential imaging.

While I do agree that the beampath configuration has a steep learning curve for new users – I feel that glossing over this aspect of the confocal use would be doing them a disservice in the long run.  I don’t expect my users to manually design beampaths every time they use the instrument – once they’ve worked out their ideal parameters the Reuse and experiment configuration functionality is invaluable – but while they are learning I feel that it is important that they understand the level of control they have over the design of their acquisition.

-Jason

-------------------------------------------------------------------
Jason M. Kirk
Technical Director, Optical Imaging & Vital Microscopy Core (OiVM)
Baylor College of Medicine
Ph: 713.798.6486
Email: [hidden email]
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[Christina Baer]

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Hi everyone,

The BINA Training and Education working group is gathering links to training resources just like these to be posted shortly as a resource on the BINA website.  If you are putting together videos or other training materials and are willing to share them more broadly, we'll be delighted to include a link to your training webpage or youtube channel.  Please don't hesitate to send links our way!  We are working on an easy submission portal, but for now, the easiest way to get us the info is via email.  

Thanks,
Christina

Christina Baer, PhD
She/her/hers
Director, SCOPE Imaging Facility
Assistant Professor
Microbiology and Physiological Systems
RNA Therapeutics Institute
Office: 508-856-6024
Cell: 650-450-7153
[hidden email]
umassmed.edu/scope
@SCOPE_UMMS

[Michelle Peckham]

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https://www.rms.org.uk/resources-downloads.html

lots of resources here

On 08/12/2020, 21:43, "Confocal Microscopy List on behalf of Christina Baer" <[hidden email] on behalf of [hidden email]> wrote:

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    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.
    *****

    Hi everyone,

    The BINA Training and Education working group is gathering links to training resources just like these to be posted shortly as a resource on the BINA website.  If you are putting together videos or other training materials and are willing to share them more broadly, we'll be delighted to include a link to your training webpage or youtube channel.  Please don't hesitate to send links our way!  We are working on an easy submission portal, but for now, the easiest way to get us the info is via email.  

    Thanks,
    Christina

    Christina Baer, PhD
    She/her/hers
    Director, SCOPE Imaging Facility
    Assistant Professor
    Microbiology and Physiological Systems
    RNA Therapeutics Institute
    Office: 508-856-6024
    Cell: 650-450-7153
    [hidden email]
    umassmed.edu/scope
    @SCOPE_UMMS

[Jason M. Kirk]

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@Dale – Smart Setup does have a ‘Smartest’ option which forces a linewise track switch.  But with more than 2 colors – it does this funky thing where it puts the most disparate fluorophores on a single track.  In my book this is a no-no unless they are very far apart – such as DAPI and AF647.  Even then I have them test controls to make sure they get no cross-talk.
 
At one time I was teaching people to use the Smart Setup to get a baseline track configuration and then modify the settings – which is a perfectly reasonable way to do things.  The reason I shied away from it was that I ended up spending more time teaching most how to modify almost all the settings manually anyway.  For our video-based references – I think it was the best play to go into the weeds immediately.

@Darran – I think it is important to keep in mind why these simplified configuration design tools like Smart Setup exist – which is to sell you a microscope.  For what they are – these tools do a pretty good job of programming rough settings for instruments that can vary wildly in their configuration.  But sadly they aren’t designed to be a rock solid configuration utility.  They are built to help vendors make their very complex instruments look like they can be operated as one operates a microwave.  This is a very slippery slope – and over time it has contributed to the idea that maybe these things should be as easy to use as a microwave.  After a swift punch in the gut though – users realize these instruments are not microwaves – and then you and I are left to work out how to help them overcome their disappointment.

@Arvydas – the emission bandwidth question is a common one and our general recommendation that I reinforce during preparations for training is that the user should be familiar with the spectral traces of their fluorophores before training (using documentation from the fluorophore vendor).  Depending on the profile they can start their collection bandwidth between 30-50nm around the central peak of the emission profile.  If the profile is broad then they can increase the bandwidth as long as that range stays away from neighboring fluorophores.