Dear all, Our lab is predominantly interested in membrane proteins and currently uses a Constant Systems device for E coli hooked up to a cooling unit which pumps 4C water around the outside of the main chamber and keeps the sample cool even when using large volumes. This works really well for E. coli but we are expressing/studying more and more (mycobacterial) proteins in M. smegmatis these days and have found that our French press is still far superior to the Constant Systems device.
The French press is aging though and I was interested if anyone else has experience with lysing M. smeg and if so which system/device they would recommend? Cheers Adam Dr Adam Heikal Department of Microbiology and Immunology University of Otago +64 (3) 471-6464 adam.hei...@otago.ac.nz http://micro.otago.ac.nz/our-people/adamheikal On 6/02/14 5:49 AM, "Michael C. Wiener" <mwie...@virginia.edu> wrote: >Interesting 'off-topic' thread. I'm a rather long-time user of >Microfluidics cell disruptors (for E. coli or P. pastoris or S. >ceresvisiae), and a shared-use M110-P Plug and Play is used by most of >the membrane-heads at my place. I've generally been happy (or at least >not unhappy) with it. > >However, we've been having some QC issues with a membrane protein that >we're making in S. ceresvisiae (Sc), and I'm having some concerns about >sample heating. Can anyone comment on Microfludics vs Avestin vs Constant >Systems vs Retsch vs whatever-else for cracking Sc cells? These days, >we're working up ~300-400g of paste at a time. > >Thank you very much! > >-MW > >Michael C. Wiener, Ph.D. >Professor >Department of Molecular Physiology >and Biological Physics >University of Virginia >PO Box 800886 >Charlottesville, VA 22908-0886 >434-243-2731 >434-982-1616 (FAX) > >On Wed, 5 Feb 2014 00:34:11 -0500 > Anirban Banerjee <ani...@gmail.com> wrote: >>I will be curious to know about people's experiences with membrane >>proteins and lysing yeast cells with the Microfluidizer and how that >>compares with using a Retsch Miller, i.e. grinding in a liquid >>nitrogen cooled stainless steel chamber and plunging in liquid >>nitrogen in between grinding cycles. >> >>I am worried that the Microfluidizer is not as mild w.r.t. heating as >>they claim it to be. That would, of course, perfectly qualify as my >>OCD. >> >>Any insights will be really appreciated. >> >>Thanks, >> >>Anirban >> >>On Tue, Feb 4, 2014 at 11:58 AM, Matthew Franklin <mfrank...@nysbc.org> >>wrote: >>> Hi Phoebe - >>> >>> "Cost-effective" may not be the applicable word here, but the >>>Microfluidizer >>> works very well: >>> >>> >>>http://www.microfluidicscorp.com/index.php?option=com_content&view=artic >>>le&id=19&Itemid=76 >>> >>> This gadget runs on house compressed air (don't try to use a >>>compressed air >>> tank - you'll empty it in minutes). It's a bit noisy, but so is a >>> sonicator. >>> >>> The Microfluidizer really shines with large volumes of lysate - like 1 >>>L and >>> up. If you're only processing 100-200 mL at a time, I think >>>sonication is >>> the best way to go. >>> >>> Hope that helps, >>> Matt >>> >>> >>> >>> On 2/4/14 11:49 AM, Phoebe A. Rice wrote: >>> >>> Some time ago, there was a nice discussion of cost-effective, wimpy >>> protein-friendly ways to break open E. coli. We're thinking about >>>replacing >>> an aging sonicator. If people have a favorite gizmo, could they >>>repeat that >>> advice? >>> thank you, >>> Phoebe Rice >>> >>> ++++++++++++++++++++++++++++++++++++++++++ >>> >>> Phoebe A. Rice >>> Dept. of Biochemistry & Molecular Biology >>> The University of Chicago >>> >>> 773 834 1723; pr...@uchicago.edu >>> http://bmb.bsd.uchicago.edu/Faculty_and_Research/ >>> >>> http://www.rsc.org/shop/books/2008/9780854042722.asp >>> >>> >>> >>> -- >>> Matthew Franklin, Ph. D. >>> Senior Scientist >>> New York Structural Biology Center >>> 89 Convent Avenue, New York, NY 10027 >>> (212) 939-0660 ext. 9374
