A new application report from Paraytec describes a new methodology using its ActiPix D100 UV area imaging detector to enable fast and label-free quantification and sizing of proteins
As well as providing superior performance over existing micro and nano flow detectors the ActiPix D100 can be set up to detect analyte plugs in multiple detection windows on the same capillary.
The new application report describes how it is possible to carry out protein quantification and sizing analyses within a capillary loop.
Using a 3nl injection of a 1mg/ml sample of bovine serum albumine (BSA, 66kDa) the UV absorption of the protein zone was recorded using two detection windows, before and after the capillary loop.
The resultant UV absorptions were fitted with HVL functions using PeakFit software to derive the area under the peaks which corresponds to the amount of protein in the sample.
The HVL parameters were used to estimate the diffusion coefficient and the size of the protein.
Unlike traditional protein sizing techniques, such as preparative agarose gel electrophoresis (Page), the ActiPix D-100 method described requires only nanolitres of the protein sample and the whole procedure can be automated.
Using the new method proteins do not require denaturation with SDS gel and can be analysed in appropriate buffers at physiological pH.
In addition using the ActiPix D-100 method there is no need for staining or labeling and further assays can be carried out following sizing directly in the same capillary.
For a copy of the new applications report and further information on the ActiPix D100 miniature UV imaging area detector please contact Paraytec using the links provided.
