A growing team of groundbreaking scientists around the world are now sharing their lab notebooks online
Over the past few weeks I’ve been perfecting a method for screening a small set of therapeutic compounds on DIPG cell lines. Before starting I made sure to discuss the techniques used in other groups to make sure that I design an assay that is comparable to theirs, although I may make my own changes Read More …
I’ve put the Molprobity stats for the model I have refined to run my fragment screening datasets against in Zenodo. It looks pretty good, and I was happy when the parameters finally all went into the green. I have about 90 datasets with resolution lower than 2.6 Å, so hopefully I’ll get at least one Read More …
As mentioned in my Zenodo post here, I fine screened ACVR1 with the three compounds in the title. I got loads of hits for co-crystallisation with K62821a. I mounted almost two pucks’ worth of crystals (31 crystals) and sent them to Diamond for Monday’s run. Unfortunately, none of them diffracted better than 3.5 Å, so Read More …
Whether or not a cell line can be transfected efficiently is an important factor to consider when designing experiments. Often times scientists would like to deliver into the cells DNA or RNA with the instructions to produce specific proteins or to delete away specific proteins. Transfection via liposome (eg: Lipofectamine 2000) is a relatively simple Read More …
Kinase activity of ALK2 and ALK5 in the cells can be quantified using specific antibodies that bind to target proteins once they are phosphorylated. However, conventional detection in Western blot requires large amount of samples and processing time (proteins are extracted from a large number of cells, linearized, separated in electric field and blotted onto a Read More …
There are concerns that compounds that are effective in inhibiting ALK2 by occupying its ATP-binding pocket might have reduced efficacy against mutant ALK2. That will be undesirable since the compounds should also target the gain-of-function mutant ALK2 in DIPG cells. The following experiment takes advantage of the fact that Activin A activates ALK2-R206H mutant but Read More …
Well, it’s been another week of not too much actual lab work and quite a lot of computer work. The lab work I did do was incredibly frustrating due to spending time searching around for reagents and then having a water problem with the liquid handler that is used to set up the fine screens. Read More …
Repeatability is a cornerstone of science, but cells behaviour can change quite significantly if you aren’t careful when looking after them. To ensure that I have the right number of DIPG cells to complete an important experiment, and that they’re all growing healthily, I want to record how fast they grow, and at what density Read More …
I haven’t done that much lab work lately as I have been working on crystal data processing among other things. However, a couple of weeks ago I purified some ACVR1/ALK2 for crystallisation for fragment screening (which happened on Friday) and for co-crystallisation with some of our compounds. The experimental details are here, on Zenodo. As Read More …
ALK2 and ALK5 are Type I receptors on the cell surface that can be activated by specific ligands (illustrated in the diagram below). Once activated by ligand, ALK2 or ALK5 interact with Type II receptor to be phosphorylated (phospho-group chemically attached to the protein). Subsequently, ALK2 phorphorylates SMAD1/5/8 while ALK5 phorphorylates SMAD2. Phosphorylated SMADs then Read More …
