Thanks for the explanation Ed, that makes sense.
----- Original Message ----- From: "Edward A. Berry" <ber...@upstate.edu> To: mi...@chem.ucla.edu Cc: CCP4BB@JISCMAIL.AC.UK Sent: Tuesday, April 5, 2011 12:22:21 PM GMT -08:00 US/Canada Pacific Subject: Re: [ccp4bb] how to Collecting Data from Long Unit Cell Axes ? When the c axis is perpendicular to the image, i.e. parallel to the xray beam, the only information about spacing along the c axis comes from the distance between the lunes. There should be a circle of spots around the direct beam with l=0, the a clear space, then a ring with l=1, then a clear space, then a ring with l=2 and so on. As the c dimension gets longer, the layers get closer together in reciprocal space, and these rings caused by intersection of the l-planes with Ewald's sphere get closer together. As the mosaicity increases, reflections whose centers are farther from ewalds sphere will be in diffraction, and the rings get broader. In the extreme of long c axis and/or high mosaicity the rings overlap completely and there appears to be only one circle of spots around the direct beam center extending out to the diffraction limit. Presumably denzo takes this to be the l=0 layer, and since it can't find the l=1 layer assumes it is out of the picture beyond the diffraction limit, which would be the case if you had an extremely short c axis so the layers are far separated. So it can get a good fit to the spot using correct a, b and a very small c. eab mi...@chem.ucla.edu wrote: > dengzq1987, > > I am also working on a data set that has this exact alignment/mosaicity > problem noted by others. If you make a movie of your dataset that displays > your images in series, or if you just click through them quickly in adxv or > your program of choice, you will see that very few of the spots ever leave > their positions on the detector as you rotate the crystal. This demonstrates > the mosaicity issue described by Bert and Ed. In my case, Denzo seems to be > able to properly measure the a and b axes of my hexagonal cell (I know this > based on a comparison of these values to structures of homologs in the same > SG), but tells me that my c axis is only 0.21A. It seems to me that in the > case of overlaps, the dimensions of the corresponding axis would be > overestimated rather than seriously underestimated (0.21A!), so I'm not > really sure of what is going on here. Maybe someone who knows more about the > software could shed some light on this crazy measurement? I've been told by a > few people that XDS may be able to handle this dataset, but I haven't gotten to trying it yet. I'll work on it today or tomorrow and follow up if I have any success. > > Best, > > Mike > > > > > > > ----- Original Message ----- > From: "Edward A. Berry"<ber...@upstate.edu> > To: CCP4BB@JISCMAIL.AC.UK > Sent: Tuesday, April 5, 2011 8:38:17 AM GMT -08:00 US/Canada Pacific > Subject: Re: [ccp4bb] how to Collecting Data from Long Unit Cell Axes ? > > In the first image (spot 2) the long unit cell is obvious from the closely > spaced > spots, but this kind of image can be handled easily by Denzo. The second one > (spot 1), although at first glance the spots seem well separated, is the > problem. > As Bert pointed out the mosaicity is high, so the lunes are overlapping > (you don't see separate rings or crescents where Ewalds sphere intersects > different layers of the lattice). That means the apparently individual > spots are multiple spots one on top of the other, with the long axis > nearly perpendicular to the plane of the picture. This means there is > no information about the spacing of the long axis in this picture,and > the picture is rather insensitive to rotation, so integration is > likely to get lost unless you fix these parameters. If you fix the > parameters based on the good orientation and integrate on through, > you will be sampling the lattice rods at intervals (as the 2D electron > crystallographers say) rather than measuring discreet spots. > However if you collect while rotating around the long axis, you will > never see this orientation and should have no serious problem. > eab > > dengzq1987 wrote: >> hello Jürgen_Bosch, >> because i have collected the data,but can't index.in some direction,the >> spot is separated .but the others are set close together(picture spot1 >> and spot 2).so we think there is one long unit cell axes. >> 2011-04-05 >> ------------------------------------------------------------------------ >> dengzq1987 >> ------------------------------------------------------------------------ >> *发件人：* Jürgen_Bosch >> *发送时间：* 2011-04-05 20:29:18 >> *收件人：* dengzq1987 >> *抄送：* CCP4BB@JISCMAIL.AC.UK >> *主题：* Re: [ccp4bb] how to Collecting Data from Long Unit Cell Axes ? >> What do you consider long ? 200, 300 ? 600 A ? Before shooting try to >> run strategy or xplan. Move the detector back to first reliably be able >> to determine your cell. Then double your estimated mosaicity and see >> what strategy suggests. If you don't get many overlaps (<5%) then try a >> closer distance. Don't rotate 1degrees but take 1/2 of the mosaicity. >> Obviously you want to make good use of the detector area so adjust the >> edges to where your crystal really diffracts. And if that resolution >> leads to too many overlaps then limit your resolution and get first a >> good datasets home. You then can play with 2theta for a higher >> resolution dataset. >> Another obvious thing to do and you don't mention what reduction program >> you use is to let XDS sort your problem out. Unless you collected to >> high resolution without being cautious XDS could help. If not, well then >> you had your experience and now should know better. >> SSRL has options to collect 450 A cells to 3A without much hassle. That >> was my largest cell so far. >> Jürgen >> >> ...................... >> 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/ >> >> On Apr 5, 2011, at 1:05, dengzq1987<dengzq1...@gmail.com >> <mailto:dengzq1...@gmail.com>> wrote: >> >>> hello all, >>> does anyone have the experience of Collecting Data from Long Unit Cell >>> Axes ? I have a crystal that diffracts to about 4 A. in some >>> /direction/ the spots overlap. we can't use the data to index .we >>> think it is because that there is a long unit cell axes. so is there >>> any method to solve this problem? >>> best wishes. >>> 2011-04-05 >>> ------------------------------------------------------------------------ >>> dengzq1987 > -- Michael C. Thompson Graduate Student Biochemistry & Molecular Biology Division Department of Chemistry & Biochemistry University of California, Los Angeles mi...@chem.ucla.edu
