Electrophoretic analysis and proteolytic activity of biological samples
We analyze the electrophoretic profile of different types of protein samples, we measure their proteolytic activity in polyacrylamide gels, and/or perform detection, quantification and typing of proteolytic activity and we determine inhibitory activity of proteases in biological samples tube.
Methodology
We can perform extraction in the laboratory from a protein extract that can be sent directly by the customer. The profile and molecular weights of the extract content shall be analysed through polyacrylamide gels. The gels will be stained with a specific stain for proteins. We use molecular weight reference markers of variable ranges according to the characteristics of the sample.
Detection of proteolytic activities in gel (zimogram) involves co-polymerization of SDS-PAGE gel with gelatin. This allows proteins with proteolytic activity to be identified, as these enzymes can degrade the gelatin present in the gel, producing clear bands on a dyed background.
The detection of tube proteases shall be determined by quantifying the azocaseinolitic activity by colorimetric method by adjusting the conditions according to the type of sample (dilution, temperature and reaction time). The activity is expressed in arbitrary units U defined as the amount of enzyme that produces an increase of A335=1 in 1 hour (A335.h-1) and is referred to by sample mL (U.mL -1) or by sample protein mg (U.mg prot. -1). Optional: serine-metallo-aspartil-cysteine protease typing; analysis by zimography. The detection of protease inhibitors will be performed pre-incubating it with a reference protease (trypsin, chymotripsine, subtilisine) and the previous assay is developed using azocasein as substrate. The result is expressed as UI inhibitor units defined as the amount of inhibitor capable of inhibiting 50% of the reference protease activity. Each determination is made in duplicate.
Implementing
Biological samples of protein nature derived from plant tissues, animals, microorganisms (for example cell suspensions and extracts, bacterial cultures, cell-free media, food, etc.).
Frequently asked questions
How to send samples?
According to the needs of the applicant, the samples can be sent refrigerated, frozen, freeze-dried, etc. You can send the proteins already extracted or we can extract them in our laboratory from a protocol that fits the characteristics of the sample and needs of the applicant.
Which companies is this service for?
The spectrum of recipients is wide. We aim at the productive sector that need to know the content, proteolytic activity and/or protease inhibitor and protein profile of their products. We have carried out services to companies linked to the food industry, such as flour production.
Technical Responsibles
Dr. Andrea Yamila Mansilla
2235186382
amansill@mdp.edu.ar
Dr. Andreina Cesari
2235134227
andreinacesari@gmail.com
Dr. María Inés Giménez
2233546671
migimen@mdp.edu.ar