Crystallization - (Mar/05/2009 )
I am completely new to crystallization. Can some one tell me how to play with lysozyme. I am aware of different methods but I would like to play with different precipitants but unfortunately I donot know how to make a recepie of of precipitants.??????
Welcome to crystalization!
You'll find lysozyme methods in any of the crystallography books and indeed in most of the catalogues. Its the standard protein to familiarise oneself with the techniques used. My seven year old came into the lab and prepared some wonderful lysozyme crystals. We now have framed colour photographs in the house of his work. Lysozyme makes beautiful crystals that are easy to shoot (x-ray) whether that be on a home source or a more powerful beam. Mind you, its difficult to justify the expense of collecting data since its been done to very small Angstroms.
Let me know if you don't find a lysozyme method. I don't have the lab book I used here with me but I'll bring it if need be.
Just had a thought. You might want to play with a plate. There are statistical concepts which I'll check up if you wish. Essentially, you make a matrix changing pH by say half a pH unit (or a whole or two units) in one direction and changing say PEG concentration in the other direction. (Need to keep really accurate notes).
Otherwise you could just try loads of different PEGs, additives and buffers. I bet the lab has loads of nearly empty bottles of reagents and buffers so you wouldn't have to make up fresh. You'll be using about 2ul per well!
If you do collect data on your lysozyme crystals, try different collection strategies. The data collection software will probably analyse the crystal and let you know what angles and how many images you need. If you then analyse the data, using the chicken wire, you can try different exclusion criteria for the spots to see the effect on the r-values, residuals etc.