16th June 2016 – Day 19

Thursday:

I feel like I have written hundreds of these blog entries now and at this point you would think I would be bored doing the same things every day but when things don’t work, which it turns out yesterday's western blot was not successful, then we get to try again making small changes as we go. 

I completed all 6 pull down assays from Monday again but this time with a lot more amylose beads, hopefully something will stick this time meaning that the western blot might actually work.

All of the samples were prepared for the four gels which we will use tomorrow, I look forward to seeing if it will work. (Cross your fingers for me.)

While preparing each sample with the help of one of my colleagues, she somehow managed to spill half a millilitre of SDS loading dye onto the bench top (if only you could have seen her face, first shock and then a naughty smile), at which point she decided to try and clean it up by pipetting it with a 10ul tip (instead of wiping it up), the two of us trying very hard not to laugh, but the tip made little dent in the little purple ‘puddle’.

The samples that the volunteers prepared from yesterday’s pull down assay were run on 3 gels and transferred to a membrane, normal WB protocol was carried out. All of the membranes were stripped of their antibodies using stripping buffer (clever name!!) and left in blocking buffer overnight. We are going to probe these tomorrow with different antibodies.

Today was a little confusing but we got there in the end.