Proteomics Platform

Protein samples can be submitted for either 1D or 2D SDS-PAGE. Gel images from Coomassie or silver-stained gels are captured, strored and analyzed using ProFinder 2-D software (Non Linear). Fluorescent gels are scanned using the ProExpress Proteomic Imaging System (PerkinElmer), an advanced fluorescence imager for the sensitive detection and quantification of low-abundance proteins in 2D gels.

Bands from 1D gels or spots from 2D gels can be cut manually or robotically using the ProXcision Proteomic Gel Cutting Robot. Profinder 2-D Image Analysis Software locates proteins of interest obtained on the ProExpress and directs the ProXcision robot to precisely cut them from the gel and place them in 96-well plates for proteolytic digestion prior to mass spectrometry. Gel pieces (bands or spots) are digested (usually with trypsin) on a MassPrep robotic workstation (PerkinElmer). The workstation can digest up to 192 samples simultaneously. Alternately, clients can submit their own gel or gel pieces. For clients running their own gels or performing gel excision, here are a few important sample preparation guidelines:

• If you run your own gels, wear gloves, lab coat, sleeves, hair net and mask during handling to avoid keratin contamination. Avoid excessive handling.

• Silver staining protocols must be compatible with mass spectrometry. Our suggested protocol is available on request. Remember that silver staining is not quantitative and it is therefore difficult to estimate the amount of protein based on the intensity of staining.

• Samples submitted for 2D gel electrophoresis must not contain SDS. Only non-ionic detergents are compatible with IEF. We can suggest IEF-friendly lysis buffers on request.

• For 2D gel electrophoresis, we require at least 50ug protein. We will desalt and measure protein concentration in your sample for a small extra charge.

• Gloves, lab coat, sleeves, hair net and a mask should be worn while cutting bands or spots for subsequent in-gel digestion. Alternately, you can bring your gel to us for cutting.

Other services include sample preparation and cleanup, immunoprecipitation, protein transfer from gels to membranes for subsequent N-terminal sequencing or western blotting.  Note that all gel-free samples submitted for LC-MS analysis will be subjected first to sample clean up.