What do you define as thin and long rods, for some of us this could be
huge crystals. What's your best guestimate regarding the size ? One
way of approximating this is taking a loop of know size and focus on
it, then focus on the crystal and guess.
Small crystals does not necessarily mean they don't diffract. I
recently got a 2.2 Å dataset with something that was 20x20 x80 µm the
largest unit cell dimension was 220 Å (needless to say the data was
collected at [advertisement on] SSRL9-2[advertisement off])
Your MPD condition will freeze no matter what you do with it, so
that's good (assuming you setup a 1:1 drop, therefore starting at 35%
MPD). How big are your drops ? If they are fairly large say 1 µl, then
the following trick might work.
A different option you could try out is the following. Premix your
protein with the 70% MPD condition in an Eppendorf tube and distribute
the mix onto a new plate. The reservoir of the plate should then have
maybe 40-60%MPD, so you are starting off closer to equilibrium but
then have a smoother gradient to the final concentration. The crystals
might take longer to appear but they might grow bigger too and you
might find some conditions without phase separation. Temperature will
definitely help but then you might need more protein.
Just some thoughts,
Jürgen
On Oct 17, 2009, at 1:44 PM, james09 pruza wrote:
Dear CCP4bbers,
Surprisingly, only At pH 3.6 the needles are appearing after 3 days
but they are very thin and long rods. I have checked it running SDS-
PAGE as well as using IZIT dye . Since the proteins has already 50mM
of NaCl so I added 100mM of NaCl in the crystallization mix. The
crystal appeared at 70% of MPD. Bunch of florets appears without
NaCl at the same condition. Will the Additives like Ethanol or 2-
propanol help in this condition? Should I not use NaCl in the mix?
Changing temperature from 4degree to 25 degree will help?
Please advice.
Thanks a lot.
James
-
Jürgen Bosch
Johns Hopkins Bloomberg School of Public Health
Department of Biochemistry & Molecular Biology
Johns Hopkins Malaria Research Institute
615 North Wolfe Street, W8708
Baltimore, MD 21205
Phone: +1-410-614-4742
Lab: +1-410-614-4894
Fax: +1-410-955-3655
http://web.mac.com/bosch_lab/
