At our last synchrotron trip, the beamline staff suggested that the problem
was due to moisture accumulation in the dry shipper. They recommended
storing them inverted (for a few weeks, if I recall), and/or putting a
supply of dry air in the dewer. Haven't tried it yet!
Nat
On Thu, Jul 11, 2013
I can't resist bashing 10.8 a little. I don't know if I would rush to
upgrade... I find the removal of 'save as' to be very annoying
(holding down option brings it back). Also the way Text edit would
hide the scroll bar by default was annoying when looking through log
files (easily changed thoug
Bei,
How do you concentrate your media? We use tangential flow filtration.
If you get a good filter (Millipore Spiral wound TFF is one example)
it goes pretty quick, in ~2 hours you should be able to process
4L(concentrate+ dilute several time in buffer). We secrete proteins
from insect cells, b
First, I don't think glycosylation occurs on lysine residues.
You should try to crystallize it as is, if that doesn't work, try to
remove the glycosylation with some or all of the following: EndoH,
PNGaseF, EndoF1, EndoF3. We have used them all to various degrees of
efficiency to remove glycans f
for 5-10L expression. For most academic
> research, there may be no need to do any viral amplification.
> Cheers,
> Chun
> Accelagen
>
> -Original Message-
> From: Nathaniel Clark
> Sender: CCP4 bulletin board
> Date: Thu, 31 Mar 20
011 at 1:12 PM, chitra shintre
wrote:
> Hi,
> We have observed this virus "going off" within 2 months as well. ...any idea
> why this happens?
> Chitra
>
> On 31 March 2011 18:07, Nathaniel Clark wrote:
>>
>> We do adherent if we have a small volume of low ti
t;>> concerning the amplification success, so we are now evaluating a new
>>> technology using qPCR with the following kit
>>> (http://oetltd.com/products/category/baculoquant/). So you might have a look
>>> and see if it could be useful for your group. We would also
We don't have a problem getting them to stick to the plates in
serum-free media, or in 5% FBS media. The more challenging part is
getting the plating density just right, too low and the plaques are
too big, to high and they are too small. Or if the cells dry out, or
if your agarose overlay is too
We use the Millipore Spiral-wound TFF1 and TFF2 cartridges for
concentrating 10-20L of insect cell media. They work great, don't
cost all that much money, and you don't need the expensive holder they
sell. You can use a ring stand and clamp the tubing directly to the
cartridge. Our cartridges ar
I use ImageJ for this purpose. There is a flag for something like
'use virtual stacks' or 'use virtual memory' that I needed to prevent
crashing.
Nat
On Mon, Mar 7, 2011 at 5:30 PM, Sean Seaver wrote:
> Dear Mark,
>
> I put together a post about creating a movie using PyMOL, eMovie (free) and
Hi,
It can, just do
fm-mode
select rmsd
I am curious though, I have heard that it is 'better' to build in
units of absolute density, but I couldn't find any values. Does any
one have a suggestion as to what absolute electron density setting is
'correct' for an Fo-Fc difference map? Or do you jus
I use a 10 ml GE-healthcare Ni-sepharose FF column, packed myself. I
strip it with EDTA after a run and then recharge before the next one.
It's has probably been used 30+ times, and I have no plans or
repacking it anytime soon, it still works as good as it ever did. We
purify secreted proteins fr
I mentioned to to Chris already, but we use nothing but HyClone
SFX-Insect powder. We make 20-30 L batches, sterilize with a large
peristaltic pump and a disposable Millipak filter from Millipore. We
never have contamination problems that are due to preparing our own
media from powder. We buy 10
Here is an incubator about the size of the Ecotherms but much cheaper:
http://www.tritechresearch.com/DT2-MP-38.html
We have one for insect cell culture that has worked great for several
years, and there is no vibration. However for crystallization we have
BOD incubators from Fisher.
Nat
On Mon
I just tried that protocol, and had essentially all of my protein
crashed out. Any tips on optimizing the methylation reaction to
reduce precipitation? Perhaps reducing the formaldehyde or
dimethylaminoborane, shortening the incubation times, etc.?
Thanks,
Nat
On Wed, Apr 14, 2010 at 6:00 PM, E
Can you soak the crystal in a solution lacking 0.2M lithium sulfate
(since it isn't your precipitant)-perhaps add 0.2M sodium chloride to
maintain ionic strength- then in a high concentration of your ligand
solution?
Nat
On Fri, Apr 23, 2010 at 3:26 PM, Jim Pflugrath wrote:
> I would test several
MiTeGen has a set of soft plastic microtools that are inteded for this
kind of stuff, and I have had suscess using them to break out single
crystals from clumps or fused crystals. You can either load in a
pencil or mount in a base.Also nylon loops. You might need to
have one tool in each hand
I have wondered if placing a layer of oil over the drop would help
solve the problem of the crystals moving around. Haven't tried it,
but don't people harvest from a microbatch tray by dragging the loop
and crystal through oil?
Nat
On Fri, Apr 9, 2010 at 11:21 AM, James Holton wrote:
> Yes, iso
Hi,
I have soaked various crystals at that pH with various sugars and
never had a problem with oxidation of the sugar. I wouldn't worry
about it. What group do you think will be oxidized?
Nat
On Fri, Mar 12, 2010 at 5:35 AM, Paul Lindblom
wrote:
> Hi,
>
> I am trying to soak some sugars in my
Jerry,
One thing you might try is a combination of Endo F1, F3, and possibly
Endo H(might trim a little bit more). This works for me on
glycoproteins expressed in insect cells which show little or no
sensitivity to PNGaseF and EndoH. Depending on the host strain you
are using to express, the sens
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