Hi Urmi, When you say "antibody" you mean Fab fragments? If so, bear in mind that Fab fragments can be quite flexible about the region inbetween the variable and constant domains, which may be detrimental to the quality of your crystals ... in this case, further to the advice of others on here, you might consider engineering an ScFv construct for your antibody. The constant domains of the antibody are not usually involved in antigen binding, so the scFv should bind to the antigen in exactly the same way as the Fab, but being less flexible it might give you better crystals. Hope that helps, Joe
> Hi, > > I am working on a protein antibody complex which readily crystallizes > (crystals form overnight and grow over 2-3 days) in 0.1M Bicine pH 9, 10 % > PEG8000. The crystals are chunky - shaped like a parallelogram but they > diffract poorly to about 8 à . > > I have tried the following to improve diffraction: > 1. Screen different temperatures 4°C - crystals have bad form and 10°C > crystals grow slower but diffraction does not improve. > 2. I have done an additive screen â A few hits came up like Yttrium > Chloride and Acetonitrile but they donât improve diffraction either > 3. I have tried streak seeding this does not help either > 4. Tested different cryo protectants â MPD, PEG400, Ethylene glycol and > glycerol - 10 - 15% glycerol seems to work best > 5. Not sure if cryo protectant affects diffraction in this case â I will > look at room temp diffraction soon to rule this out. > 6. Typical diffraction images attached > > Does anyone have suggestions on what I could try to improve diffraction of > my crystals? > > > > Urmi Dhagat > St Vincent's Institute > > >
