[Xenopus] [External] Re: MISC
Ira Blitz
ilblitz at uci.edu
Thu Oct 29 11:59:24 EDT 2020
Anna,
I was just going to email to recommend Jonathan Slack’s trypsin method but he beat me to it, so I’ll just send this to say it works as advertised as I’ve tried it!
Ira
> On Oct 29, 2020, at 7:21 AM, slack017 University of Minnesota <slack017 at umn.edu> wrote:
>
> Hi Anna et al.
> In my day there was no Gastromaster and we cut loads of animal caps with sharpened tungsten needles and a hair loop or forceps to steady the embryo. To separate layers in neurulae it helps a lot to include a little trypsin in the medium. Leave the Ca in then you tend to degrade the ECM but not to disaggregate the cells. Methods are in the attached (otherwise uninteresting) paper.
> Good luck,
> Jonathan
>
> Jonathan M.W.Slack, PhD, FMedSci.,
> Emeritus Professor, University of Bath, UK.
> Emeritus Professor, University of Minnesota, USA.
> http://www.bath.ac.uk/bio-sci/contacts/academics/jonathan_slack/ <http://www.bath.ac.uk/bio-sci/contacts/academics/jonathan_slack/>
> On Thu, Oct 29, 2020 at 1:27 PM Klein, Peter <pklein at pennmedicine.upenn.edu <mailto:pklein at pennmedicine.upenn.edu>> wrote:
> Hi Leon,
>
>
>
> The gastromaster is much faster but in my lab only works with the younger generation of scientists, who could easily cut 50 or more in an hour. Either approach takes a little practice, but given shortage of tips and lack of information about how to make them, I agree with above: you could cut hundreds with a good team, some #5 Dumonts, and hair knives.
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> Peter
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> From: <xenopus-bounces at lists.mbl.edu <mailto:xenopus-bounces at lists.mbl.edu>> on behalf of Anna Philpott <ap113 at cam.ac.uk <mailto:ap113 at cam.ac.uk>>
> Date: Thursday, October 29, 2020 at 8:48 AM
> To: "Green, Jeremy" <jeremy.green at kcl.ac.uk <mailto:jeremy.green at kcl.ac.uk>>, Leon Peshkin <peshkin at gmail.com <mailto:peshkin at gmail.com>>
> Cc: "xenopus at lists.mbl.edu <mailto:xenopus at lists.mbl.edu>" <xenopus at lists.mbl.edu <mailto:xenopus at lists.mbl.edu>>
> Subject: [External] Re: [Xenopus] MISC
>
>
>
> Dear Leon,
>
> OMG I would LOVE a cap cutting jamboree. We have a gastromaster on the shelf with some tips but also found it less than the hype and as we can’t get tips any more and they break a lot, it’s not much of a solution. Yes, 1 also think No5s and some experience are the only way forward-freezing in batches if necessary for pooling later. Do you need caps rather than animal poles? Razor blades are quicker obvs. We are currently trying to compare ecto/meso with endoderm. This unfortunately means peeling the epidermis off the ventral tissue (neurula stages) to get pure endoderm. This is agonising. Does anyone have an easy way of doing that?
>
> Anna
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> Prof. Anna Philpott
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> Head of the School of Biological Sciences
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> And
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> Group Leader, Cambridge Stem Cell Institute
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> University of Cambridge
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> From: <xenopus-bounces at lists.mbl.edu <mailto:xenopus-bounces at lists.mbl.edu>> on behalf of Jeremy Green <jeremy.green at kcl.ac.uk <mailto:jeremy.green at kcl.ac.uk>>
> Date: Thursday, 29 October 2020 at 11:13
> To: Leon Peshkin <peshkin at gmail.com <mailto:peshkin at gmail.com>>
> Cc: "xenopus at lists.mbl.edu <mailto:xenopus at lists.mbl.edu>" <xenopus at lists.mbl.edu <mailto:xenopus at lists.mbl.edu>>
> Subject: Re: [Xenopus] MISC
>
>
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> Dear Leon,
>
>
>
> With a bit of practice and well-sharpened Dumont #5 forceps, a person can cut around 200 caps in an hour. I suggest that if you want to get many hundreds of caps that you simply have a cap-cutting jamboree - not so much biting the bullet as having a party. Seriously. I think you’ll have no problem getting volunteers and (COVID, etc. permitting), I’d be happy to come over myself.
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> Best wishes,
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> Jeremy
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> Jeremy B.A. Green Ph.D.
>
> Vice-Dean International (Research)
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> Professor of Developmental Biology
>
> Centre for Craniofacial & Regenerative Biology
>
> King's College London
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> Guy's Tower, Floor 27
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> London SE1 9RT
>
> UK
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>
> jeremy.green at kcl.ac.uk <mailto:jeremy.green at kcl.ac.uk>
> Tel. +44 20 7188 1794
> https://www.kcl.ac.uk/people/jeremy-green <https://www.kcl.ac.uk/people/jeremy-green>
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> On 29 Oct 2020, at 03:45, Leon Peshkin <peshkin at gmail.com <mailto:peshkin at gmail.com>> wrote:
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> Dear Colleagues !
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> I have a few general questions and tried posting to Slack "Xenopus Community" without success. People - Slack is a nice way to keep discussions going, please use it !
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> Has someone worked out a way to make animal caps en masse ? I did hear the gastromaster legend and even obtained one with a few tips … but it does not really work in my hands. Any other devices ? Please do not tell me to just bite the bullet. I'd like to be able to cut many hundreds.
>
> There are a couple of animal cap instructional videos with forceps but I know many people use other techniques. Would be nice to have more videos.
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> Does someone have a protocol and experience for blood perfusion in Xenopus ?
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> THANK YOU
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> - L. Peshkin
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