So, yeah, my name is Rhys, and my project is developing molecular standards for the

AUC for the use in the CGMP environment.

So, quickly give a breakdown of the work I've completed so far and my plan for the

future.

So, of course, molecular standards play a very important role in the biopharma industry.

They ensure that the strict quality control measures are adhered to.

So in developing my molecular standards based upon double-stranded DNA molecules, we hope

to provide a way for AUC instruments to be validated in a current good manufacturing

practices space.

So you may be asking, well, why do we need molecular standards for the AUC, right?

AUC is the first principles technique.

As long as our rotor speed, temperature, optical system, and radio positioning system are calibrated

correctly, we should get the correct answers all the time.

However, any imprecise calibrations in this are not so easily detected simply based upon

the data.

So, a validated standard would provide value in especially comparing from lab to lab and

instrument to instrument.

So not going to go too much in depth here since this is not particularly relevant, but

using purifying and getting my fragments ready for the AUC, we do this using this recombinantly

produced plasmid.

So we produce two different size fragments, one 2,000H11-base pairs long, the other 208-base

pairs long.

Purify this, of course, using size exclusion chromatography.

We can then confirm this, of course, using agarose gel electrophoresis.

Following this, we're, of course, ready to stick these in the AUC.

So we take our two different size fragments as well as some mixtures of them.

We came up with a variety of rotor speeds and temperatures to kind of reflect a broad

range of experimental conditions that may be used.

We used the adaptive space time finite element method simulator in Ultrascan to guide our

experimental design in this case.

And of course, we stick those into the AUC and we hope to get back reproducible values

as well as expected variations in sedimentation coefficient, diffusion coefficient, and frictional

ratio.

So quickly some of the data that I've collected so far on our smaller fragment and then on

our larger fragment here.

So fitting globally for the 208-base pair fragment across 30 kilorpms and 50 kilorpms,

we have nice, clean, tight distributions for our fragments here.

And of course, we have our van hold Weische plots showing nice homogeneous boundaries,

especially at 20 degrees Celsius and 37 degrees Celsius.

And then we have a table tabulating the data for sedimentation coefficient, diffusion coefficient,

and the frictional ratio.

Of course, this is only from one replicate of the data.

So of course, we need to collect a lot more data to establish expected variations in these

parameters.

And then for the larger fragment, once again, nice, tight discrete sedimentation coefficient

distributions based upon the global fit across those three different rotor speeds at each

temperature.

Our van hold Weische distributions once again for each of the speeds.

And then the tabulated results for SD and the frictional ratio.

So another important component of our study is of course understanding how long the molecular