 |
 |
Thank you for your comments everyone. The CCP4BB is a wonderful resource and it has answered several questions that have been bothering me for years! Tristran has given us the correct conclusion as well as the important facts: the capacity of oil for holding O2 is high, but the diffusion rate is low. This makes complete sense of the observations reported. The O2 is (slowly) diffusing OUT of the permanganate drop, and the oil is already saturated with O2, therefore it takes a long time for the purple colour to be lost. The O2 is diffusing INTO the drop in the dithionite experiment, and presumably the oil that Julia used was already loaded with O2, so the reducing environment was quickly lost. I hadn't figured out how to take advantage of the protection of oil - I just had a vague feeling that it might be helpful. Now however I can see that it's useful, because the oil will provide quite good protection against a pulse of O2 e.g. if someone accidentally lets air into the chamber. (Or moves plates from one glovebox to another?) O2 will start to diffuse into the oil, but most of it will diffuse out again if the O2 pulse is short. And the lids that are standard on microbatch plates will help a lot. (The oil is almost the ideal barrier, although you *might* prefer something with a very low solubility since it *might *give a lower O2 flux in the steady state - as Tristran says it's complicated. And I'm guessing that the O2 flux through the thin plastic tape used in vapor diffusion setups would be quite high. Does anyone have a friend who works in food science?) There's an even more exciting conclusion: we should degass our oil even for use with *aerobic *microbatch setups. I have heard of a case where diffracting crystals were only obtained for aerobic targets in a glovebox, and I think the skins on drops are, or can be, related to oxidation. There may even be mileage in microbatch with the zip lock bag approach for targets that are not overly sensitive - *if* you degas your oil before you start .... a vacuum should do it. I agree that Al's Oil (silicone) should be avoided from this point of view - although I would certainly use it anyway for screening experiments (whether aerobic or anaerobic). Riveting stuff. Thx to all, Patrick On 18 March 2015 at 18:58, Tristan Croll <[log in to unmask]> wrote: > It's a little complicated. It's true that oxygen is more soluble in most > oils than in water - but in a high viscosity mineral oil the diffusion rate > is orders of magnitude lower. So the combination of an oil overlay and a > reducing agent in your buffer should protect your sample much longer than > the reducing agent alone - as long as your oil was degassed to start with. > Note that silicon oils are a bad choice for this - silicones have an > enormous affinity for oxygen (so much so that they've been explored as > artificial blood substitutes), and it diffuses through them very readily. > > > > Tristan Croll > Lecturer > Faculty of Health > School of Biomedical Sciences > Institute of Health and Biomedical Engineering > Queensland University of Technology > 60 Musk Ave > Kelvin Grove QLD 4059 Australia > +61 7 3138 6443 > > This email and its attachments (if any) contain confidential information > intended for use by the addressee and may be privileged. We do not waive > any confidentiality, privilege or copyright associated with the email or > the attachments. If you are not the intended addressee, you must not use, > transmit, disclose or copy the email or any attachments. If you receive > this email by mistake, please notify the sender immediately and delete the > original email. > > > > > On 18 Mar 2015, at 11:49 pm, Edward A. Berry <[log in to unmask]> wrote: > > > > Do you have evidence that the oil blocks diffusion of O2? O2 is a > nonpolar molecule, generally much more soluble in oils than in water. I'm > not sure about silicone oils, but I would think they also dissolve O2 > readily. > > eab > > > >> On 03/18/2015 08:02 AM, Patrick Shaw Stewart wrote: > >> > >> Hi Steve > >> > >> I have one more comment for this thread. > >> > >> The microbatch-under-oil method is very handy for anaerobic work: > >> > >> 1. You can keep the microbatch stock solutions in normal microtitre > plates (polypropylene is best to reduce evaporation) for months, which > hugely reduces the amount of degassing that you need to do. You will only > use say 0.5 ul of stock per drop. > >> > >> 2. The oil offers a surprising amount of protection from oxidation, > which may be helpful eg in harvesting. > >> > >> 3. Microbatch can be automated - in parallel to vapor diffusion if > desired > >> > >> > >> It's amazing how often (aerobic) microbatch produces far superior > crystals to V.D. for no obvious reason - it's well worth trying for both > screening and optimization. > >> > >> Best wishes > >> > >> Patrick > >> > >> > >> > >> On 11 March 2015 at 10:17, <Stephen Carr> < > [log in to unmask] <mailto:[log in to unmask]>> > wrote: > >> > >> Dear CCP4BBer's > >> > >> Apologies for the off-topic post, but the CCP4BB seems to be the > best place to ask about crystallisation. > >> > >> I am looking to set up crystallisation in an anaerobic glove box and > wondered how other people did this, specifically the crystallisation > stage. My initial thoughts were to place a small crystallisation incubator > inside the box, however the smallest I have come across so far (~27L) is > still rather large. Has anyone come across smaller incubators? > Alternatively are incubators even neccessary if the glove box is placed in > a room with good air conditioning and stable temperature control? > >> > >> Any recommendations would be very helpful. > >> > >> Thanks in advance, > >> > >> Steve Carr > >> > >> Dr Stephen Carr > >> Research Complex at Harwell (RCaH) > >> Rutherford Appleton Laboratory > >> Harwell Oxford > >> Didcot > >> Oxon OX11 0FA > >> United Kingdom > >> Email [log in to unmask] <mailto: > [log in to unmask]> > >> tel 01235 567717 <tel:01235%20567717> > >> > >> This email and any attachments may contain confidential, copyright > and or privileged material, and are for the use of the intended addressee > only. If you are not the intended addressee or an authorized recipient of > the addressee, please notify us of receipt by returning the e-mail and do > not use, copy, retain, distribute or disclose the information in or > attached to this email. > >> > >> Any views or opinions presented are solely those of the author and > do not necessarily represent those of the Research Complex at Harwell. > >> > >> There is no guarantee that this email or any attachments are free > from viruses and we cannot accept liability for any damage which you may > sustain as a result of software viruses which may be transmitted in or with > the message. > >> > >> We use an electronic filing system. Please send electronic versions > of documents, unless paper is specifically requested. > >> > >> This email may have a protective marking, for an explanation, please > see: > >> > http://www.mrc.ac.uk/About/informationandstandards/documentmarking/index.htm > . > >> > >> > >> > >> > >> -- > >> [log in to unmask] <mailto:[log in to unmask]> Douglas > Instruments Ltd. > >> Douglas House, East Garston, Hungerford, Berkshire, RG17 7HD, UK > >> Directors: Peter Baldock, Patrick Shaw Stewart > >> > >> http://www.douglas.co.uk > >> Tel: 44 (0) 148-864-9090 US toll-free 1-877-225-2034 > >> Regd. England 2177994, VAT Reg. GB 480 7371 36 > -- [log in to unmask] Douglas Instruments Ltd. Douglas House, East Garston, Hungerford, Berkshire, RG17 7HD, UK Directors: Peter Baldock, Patrick Shaw Stewart http://www.douglas.co.uk Tel: 44 (0) 148-864-9090 US toll-free 1-877-225-2034 Regd. England 2177994, VAT Reg. GB 480 7371 36