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Gel Based Proteomics

Gel electrophoresis is an important methodology employed for protein analysis, which contains many different techniques, including sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), isoelectric focusing (IEF), and so on. SDS-PAGE, the most common gel electrophoresis used for proteins, provides an easy method to estimate the molecular weight of proteins, assess the complexity of the sample or the purity of a preparation, and monitor the fractions obtained during chromatographic or other purification procedures.

 

PMF (Peptide Mass Fingerprinting)

 

GenePrint LifeSciences offers Peptide Mass Fingerprinting (PMF), which is an analytical technique for identifying unknown proteins. Proteins to be identified are first digested into peptides, whose absolute mass can be accurately analyzed by a mass spectrometer, such as MALDI-TOF or ESI-TOF. MALDI is more preferred in PMF analysis, as it has a wide sequence coverage and can analyze a large amount of sample at the same time. The peptide mass are compared to database containing the theoretical peptide masses of all known protein sequences. The match with the highest score is considered to be the identity of the unknown protein. As only the masses of peptide, not the exact sequence of peptides, are measured, there can some interference to the mass match. For example, post-translational modification results in up-shift of the peptide mass, and contamination of other protein also affect the accuracy of the analysis. Therefore, large portion of post-translational modifications and high contamination of other proteins should be eliminated before the analysis.

 

DIGE

 

One of the largest challenges in proteomics today is to be able to quantify the composition and amount of proteins found in a specific cell or tissue at a defined time point. GenePrint LifeSciences offers differential gel electrophoresis (DIGE), which is a gel electrophoresis‚Äźbased technique for protein quantification in complex mixtures. In DIGE, the high resolution of two‚Äźdimensional gel electrophoresis is combined with the excellent dynamic range obtained by fluorescent tag labelling of protein samples. The output of DIGE experiments provides information about how many proteins display changed expression levels on a specific treatment. In addition, proteins of interest can be excised and identified with conventional mass spectrometry techniques and further analysed by other biochemical methods.

 

1D/ SDS-PAGE

1D SDS-PAGE is a method that separates proteins by molecular weight, where the samples are weighed and dissolved in SDS. GenePrint LifeSciences offers SDS-PAGE, which is widely used in protein analysis, such as protein size determination, protein identification, sample purity analysis, and so on. IEF is an electrophoretic technique to separate proteins based on isoelectric point (pI), which can resolve different proteins of similar size that cannot be resolved on standard SDS-PAGE gels.

 

2D Gel Electrophoresis

 

GenePrint LifeSciences offers two dimensional gel electrophoresis. 2-DE is a powerful tool for separation and fractionation of complex protein mixtures. The 2-DE separates protein firstly by IEF and then by SDS-PAGE. The isoelectric point is defined as the pH at which the net charge of the protein becomes zero. Isoelectric point analysis can be used to study heterogeneity, analyze protein-folding status, and predict protein-protein interactions. High-resolution 2D electrophoresis is used for protein species and isoform distribution analysis, host cell protein analysis, quantitative comparison of cell states, and so on.

 

Western Blot and Electrical Transfer Analysis

GenePrint LifeSciences offers Western blot, which is a commonly used method for protein analysis, such as qualitative and semi-quantitative analysis. In this method, proteins are separated by 2-DE and then these proteins are transferred to a carrier membrane. Western blot can be used for the qualitative detection of single proteins and protein modifications.

 

2D Blue Native / SDS-PAGE for Complex Analysis

GenePrint LifeSciences offers blue native (BN) PAGE, one of the native gel methods, which can analyze proteins that are in a folded state. In 2D BN/SDS-PAGE analysis, proteins are analyzed by dimension BN-PAGE and second dimension SDS-PAGE. 2D BN/SDS-PAGE has been proven to be a useful tool for protein complex analysis. It can provide information about the size, number, protein composition, stoichiometry and relative abundance of the sample. GenePrint LifeSciences provides a method for the global analysis of the subunits of protein complexes through 2D Blue Native / SDS-PAGE.

 

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