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Comparative evaluation of plasma protein purification and 2-D gel electrophoresis protocols for analysis of HIV-1 infected human plasma proteins

Barik, S. K.; Barik, D. K.; Mohanty, K. K.; Bisht, D.; Patil, S. A.; Singh, R.; Sharma, D. K.; Tripathy, S. P.; Tandon, R.; Singh, T. P.; Jena, S.

2021-03-04 biochemistry
10.1101/2021.03.03.433830 bioRxiv
Show abstract

Purification of proteins from human plasma is a herculean task to perform 2-D gel electrophoresis. Human plasma contains nearly 70% albumin and globulin. The removal of such high abundance high molecular weight proteins is very difficult before performing 2-D gel electrophoresis. It becomes more difficult when we intent to investigate in infectious diseases like HIV/AIDS. We tried to the best of our efforts adopting various organic and non-organic based protocols based on various published papers. After failure of these protocols in results of 2-D gel-electrophoresis Aurum serum mini kit (Bio-Rad, USA) was adopted for plasma protein purification for performing 2-D gel electrophoresis. The low-abundance proteins were better resolved by 10% SDS-PAGE in 2-D gel-electrophoresis. Then,we extended the MALDI-TOF/TOF analysis of low-abundance proteins in human plasma by adopting the Aurum serum mini kit (Bio-Rad, USA) for 2-D gel electrophoresis. Thus, we concluded that, depletion of high abundant proteins like albumin and globulin, the use of the Aurum serum mini kit (Bio-Rad, USA) is the protocol of choice to perform the 2-D gel electrophoresis of HIV-1 infected human plasma.

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