Mosflm was originally developed by Andrew G.W. Leslie and is now being updated by Harry Powell at MRC Laboratory of Molecular Biology.
Mosflm can process diffraction images from a wide range of detectors and produces, as output, an MTZ file of reflection indices with their intensities and standard deviations (and other parameters). This MTZ file is passed onto other programs of the CCP4 program suite (SORTMTZ, SCALA, TRUNCATE) for further data reduction (reference).
In addition, Harry has a page dedicated toward help and guidance using mosflm which can be found here.
To get started using mosflm (assuming mosflm has already been installed) you have to open a terminal then type ‘ipmosflm’ or ‘imosflm’ (you don’t need a comm file if you are using iMosflm) depending on which version you would like to use. Here is a list of the basic linux commands that you will need to use the program.
This information is also available in the MOSFLM 7.0.4 User Guide. My goal here is to have a comm file that you can simply copy/paste/edit. A “comm file” is a file that allows mosflm to understand inputs specific to your experiment that may vary between users (such as what you named your files or where they are located).
A comm file is simply a text file, which can be generated by typing ‘edit’ into your terminal. Once completed save this file in the directory in which you plan to run mosflm. Save the file with the name ‘comm’ so it is easy to remember when you need to refer to the document using mosflm.
If you would like more information about creating a comm file, you may find this link help.
Below is a hypothical example of what your comm file should contain:
detector marccd (type of dectector used during collection)
findspots threshold 15 (threshold to search for spots this is a good default, but can adjust later)
synchrotron polarisation 1.00 (wavelength used during collection)
directory /home/data/lyso (location of the data)
template lyso_### (what the data files are named)
image 1 phi 0 0.3 (the number of the 1st image, usually 1 and the step size along phi in this case)
The # symbols stand for any number which allows all your images to be read into mosflm.
This is a blank comm file that you will need to fill with your experimental data:
detector
findspots threshold
synchrotron polarisation
directory
template
image 1 phi
You may also need other inputs depending on if your image header was correctly formatted which would include:
distance 120.00
beam 84.444 84.824
pixel 0.0792
How do I prevent condensation on cover slips?
Condensation on your cover slips can easily ruin your crystallization setup. The fact that this may occur after a number days/weeks/months of protein preparation can make this event especially aggravating.
I recently came across a number of posts by Partrick Stewart from Douglas Instruments. I have combined and modified the posts and hopefully provided a sensible thermodynamic explanation. Finally, I have also included a tip to prevent condensation from occurring.
1. If you have condensation, then you MUST have a heat flow, where the heat is flowing to where the condensation occurs.
2. The solutes in the reservoir will stop condensation due to minor heat flows. The more salt etc, the less condensation on the tape.
I find the easy way to think about condensation is to remember that wherever there is a heat flow, that flow carries moisture with it. (Or you could think of it as the moisture carries the heat, although obviously it is the heat flow that drives the process.)
Therefore, when you put a warm plate in a cold-room, heat will flow UPWARDS from the reservoirs into the air above the plate, and you’ll get condensation on the tape or cover slip.
The simple solution is to put a (warm) book from your office on top of the plate when you put it into the cold room. Now you have a heat flow DOWNWARDS from the book, through the plate and into the cold bench.
This method is guaranteed to prevent condensation – in fact it will remove condensation if you have it – just put a warm book on the plate.
I haven’t experimented with the size of book – it could be that I have been actively drying out the drops, which may not be so good!
And I’m sure that other objects would work well although I have only tried books. They seem ideal because they have a reasonable thermal mass but quite low thermal conductivity.
Finally, you will have a use for those Harry Potter books.
Macromolecular crystallography is a biophysical technique used to understand biological molecules such as proteins, viruses, RNA and DNA to near atomic resolution. This high resolution helps scientist understand the mechanism by which these macromolecules carry out their functions in living cells and organisms.
Macromolecules can be generated from recombinant technology, synthetically developed or from a natural source, all of which require purification. Macromolecules can crystallize under the right conditions to form repeating units in a regular 3 dimensional lattice. Once crystals have been achieved, X rays can then be diffracted by the atoms in equivalent positions in the crystal lattice that results in sharp intense spots (called a diffraction pattern). The macromolecular structure can be determined by the analysis of the intensities and positions of the diffraction spots.