Drug Delivery and Translational Research

, Volume 3, Issue 5, pp 428–436 | Cite as

Optimized lysis buffer reagents for solubilization and preservation of proteins from cells and tissues

  • Byeong Hee Hwang
  • Kenneth Y. Tsai
  • Samir Mitragotri
Research Article

Abstract

Reagents that facilitate solubilization of cells and tissues while preserving the biological activity of their constituents play a major role in various applications including drug delivery. Such reagents are necessary for the accurate determination of cellular and tissue concentrations of proteins, peptides, and nucleic acids, and to measure therapeutic efficacy of drug delivery technologies. Surfactant-based reagents are commonly used for this purpose; however, their utility is marred either by limited ability to solubilize or tendency to denature the proteins during solubilization. Here, we report on the screening and identification of combinations of nonionic and zwitterionic surfactants that possess excellent ability to solubilize mechanically strong and elastic tissues such as skin, while preserving its protein constituents. The leading combination, comprising an equi-mass mixture of 3-(N,N-dimethyl myristyl ammonio) propanesulfonate (TPS, CAS number:14933-09-6) and polyoxyethylene(10) cetyl ether (Brij® C10, CAS number: 9004-95-9) with a total surfactant concentration 0.5 % w/v, solubilized keratinocytes and preserved the activity of glyceraldehyde 3-phosphate dehydrogenase (GAPDH) enzyme in its extracts at room temperature for 7 days. The ability of this mixture to preserve GAPDH activity far exceeded that of a commonly used reagent, Triton-X100. The same mixture also helped solubilize mouse skin to extract proteins and maintain detectable activity of GAPDH in the extract for 1 day. Several other mixtures of nonionic and zwitterionic surfactants were studied. These mixtures provide new reagents for solubilization of cells and tissues for research as well as technological applications.

Keywords

Reagent Solubilization Preservation Surfactant Detergent Assay 

References

  1. 1.
    Hosfield D, Palan J, Hilgers M, Scheibe D, McRee DE, Stevens RC. A fully integrated protein crystallization platform for small-molecule drug discovery. J Struct Biol. 2003;142(1):207–17.PubMedCrossRefGoogle Scholar
  2. 2.
    Oellerich M, Barten MJ, Armstrong VW. Biomarkers—the link between therapeutic drug monitoring and pharmacodynamics. Therapeutic Drug Monitoring. 2006;28(1):35–8.PubMedCrossRefGoogle Scholar
  3. 3.
    Arrell DK, Niederlander NJ, Perez-Terzic C, Chung S, Behfar A, Terzic A. Pharmacoproteomics: advancing the efficacy and safety of regenerative therapeutics. Clin Pharmacol Ther. 2007;82(3):316–9.PubMedCrossRefGoogle Scholar
  4. 4.
    Futcher B, Latter GI, Monardo P, McLaughlin CS, Garrels JI. A sampling of the yeast proteome. Mol Cell Biol. 1999;19(11):7357–68.PubMedGoogle Scholar
  5. 5.
    VerBerkmoes NC, Denef VJ, Hettich RL, Banfield JF. Systems biology: functional analysis of natural microbial consortia using community proteomics. Nat Rev Microbiol. 2009;7(3):196–205.PubMedCrossRefGoogle Scholar
  6. 6.
    Paulo CS, Pires das Neves R, Ferreira LS. Nanoparticles for intracellular-targeted drug delivery. Nanotechnology. 2011;22(49):494002.PubMedCrossRefGoogle Scholar
  7. 7.
    Ariga K, McShane M, Lvov YM, Ji Q, Hill JP. Layer-by-layer assembly for drug delivery and related applications. Expert Opin Drug Deliv. 2011;8(5):633–44.PubMedCrossRefGoogle Scholar
  8. 8.
    De Cock LJ, De Koker S, De Geest BG, Grooten J, Vervaet C, Remon JP, et al. Polymeric multilayer capsules in drug delivery. Angew Chem Int Ed Engl. 2010;49(39):6954–73.PubMedCrossRefGoogle Scholar
  9. 9.
    Cosgrove BD, Alexopoulos LG, Hang TC, Hendriks BS, Sorger PK, Griffith LG, et al. Cytokine-associated drug toxicity in human hepatocytes is associated with signaling network dysregulation. Mol Biosyst. 2010;6(7):1195–206.PubMedCrossRefGoogle Scholar
  10. 10.
    Fournier E, Passirani C, Montero-Menei CN, Benoit JP. Biocompatibility of implantable synthetic polymeric drug carriers: focus on brain biocompatibility. Biomaterials. 2003;24(19):3311–31.PubMedCrossRefGoogle Scholar
  11. 11.
    Scopes RK, Cantor CR, editors. Protein purification: principles and practice. Springer: New York; 1994. p. 22–43.Google Scholar
  12. 12.
    Heredia KL, Bontempo D, Ly T, Byers JT, Halstenberg S, Maynard HD. In situ preparation of protein-"smart" polymer conjugates with retention of bioactivity. J Am Chem Soc. 2005;127(48):16955–60.PubMedCrossRefGoogle Scholar
  13. 13.
    Brown RB, Audet J. Current techniques for single-cell lysis. J R Soc Interface. 2008;5(supp 2):S131–8.PubMedCrossRefGoogle Scholar
  14. 14.
    Paliwal S, Ogura M, Mitragotri S. One-step acquisition of functional biomolecules from tissues. Proc Natl Acad Sci U S A. 2010;107(33):14627–32.PubMedCrossRefGoogle Scholar
  15. 15.
    Hwang B, Doshi N, Tsai KY, Mitragotri S. A reagent to facilitate protein recovery from cells and tissues. Drug Delivery Translational Res. 2012;2(5):297–304.CrossRefGoogle Scholar
  16. 16.
    Mitragotri S, Ray D, Farrell J, Tang H, Yu B, Kost J, et al. Synergistic effect of low-frequency ultrasound and sodium lauryl sulfate on transdermal transport. J Pharm Sci. 2000;89(7):892–900.PubMedCrossRefGoogle Scholar
  17. 17.
    Tezel A, Sens A, Tuchscherer J, Mitragotri S. Synergistic effect of low-frequency ultrasound and surfactants on skin permeability. J Pharm Sci. 2002;91(1):91–100.PubMedCrossRefGoogle Scholar
  18. 18.
    Fekete M, Wittliff JL, Schally AV. Characteristics and distribution of receptors for [D-TRP6]-luteinizing hormone-releasing hormone, somatostatin, epidermal growth factor, and sex steroids in 500 biopsy samples of human breast cancer. J Clin Lab Anal. 1989;3(3):137–47.PubMedCrossRefGoogle Scholar
  19. 19.
    Anson ML. The denaturation of proteins by synthetic detergents and bile salts. J Gen Physiol. 1939;23(2):239–46.PubMedCrossRefGoogle Scholar
  20. 20.
    Daskal I, Ramirez SA, Ballal RN, Spohn WH, Wu B, Busch H. Detergent lysis for isolation of intact polysomes of Nivikoff hepatoma ascites cells. Cancer Res. 1976;36(3):1026–34.PubMedGoogle Scholar
  21. 21.
    Rabilloud T, Adessi C, Giraudel A, Lunardi J. Improvement of the solubilization of proteins in two-dimensional electrophoresis with immobilized pH gradients. Electrophoresis. 1997;18(3–4):307–16.PubMedCrossRefGoogle Scholar
  22. 22.
    Cox RA. The use of guanidinium chloride in the isolation of nucleic acids. Methods Enzymology. 1968;12:120–9.CrossRefGoogle Scholar
  23. 23.
    O'Farrell PH. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975;250(10):4007–21.PubMedGoogle Scholar
  24. 24.
    Ullrich A, Shine J, Chirgwin J, Pictet R, Tischer E, Rutter WJ, et al. Rat insulin genes: construction of plasmids containing the coding sequences. Science. 1977;196(4296):1313–9.PubMedCrossRefGoogle Scholar
  25. 25.
    Chomczynski P, Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987;162(1):156–9.PubMedCrossRefGoogle Scholar
  26. 26.
    Tan SC, Yiap BC. DNA, RNA, and protein extraction: the past and the present. J Biomed Biotechnol. 2009;2009:574398.PubMedCrossRefGoogle Scholar
  27. 27.
    Rigaut G, Shevchenko A, Rutz B, Wilm M, Mann M, Seraphin B. A generic protein purification method for protein complex characterization and proteome exploration. Nat Biotechnol. 1999;17(10):1030–2.PubMedCrossRefGoogle Scholar
  28. 28.
    Ho Y, Gruhler A, Heilbut A, Bader GD, Moore L, Adams SL, et al. Systematic identification of protein complexes in Saccharomyces cerevisiae by mass spectrometry. Nature. 2002;415(6868):180–3.PubMedCrossRefGoogle Scholar
  29. 29.
    Ge H. UPA, a universal protein array system for quantitative detection of protein–protein, protein–DNA, protein–RNA and protein–ligand interactions. Nucleic Acids Res. 2000;28(2):e3.PubMedCrossRefGoogle Scholar
  30. 30.
    Wiedenmann B, Franke WW. Identification and localization of synaptophysin, an integral membrane glycoprotein of Mr 38,000 characteristic of presynaptic vesicles. Cell. 1985;41(3):1017–28.PubMedCrossRefGoogle Scholar
  31. 31.
    Paliwal S, Hwang BH, Tsai KY, Mitragotri S. Diagnostic opportunities based on skin biomarkers. Eur J Pharm Sci. 2012. doi:10.1016/j.ejps.2012.10.009.
  32. 32.
    Cho SW, Goldberg M, Son SM, Xu QB, Yang F, Mei Y, et al. Lipid-like nanoparticles for small interfering RNA delivery to endothelial cells. Adv Funct Mater. 2009;19(19):3112–8.PubMedCrossRefGoogle Scholar
  33. 33.
    Kuroda S, Yamazaki M, Abe M, Sakimura K, Takayanagi H, Iwai Y. Basic leucine zipper transcription factor, ATF-like (BATF) regulates epigenetically and energetically effector CD8 T-cell differentiation via Sirt1 expression. Proc Natl Acad Sci U S A. 2011;108(36):14885–9.PubMedCrossRefGoogle Scholar
  34. 34.
    Agbor TA, Cheong A, Comerford KM, Scholz CC, Bruning U, Clarke A, et al. Small ubiquitin-related modifier (SUMO)-1 promotes glycolysis in hypoxia. J Biol Chem. 2011;286(6):4718–26.PubMedCrossRefGoogle Scholar
  35. 35.
    Durrieu C, Bernier-Valentin F, Rousset B. Microtubules bind glyceraldehyde 3-phosphate dehydrogenase and modulate its enzyme activity and quaternary structure. Arch Biochem Biophys. 1987;252(1):32–40.PubMedCrossRefGoogle Scholar
  36. 36.
    Park J, Han D, Kim K, Kang Y, Kim Y. O-GlcNAcylation disrupts glyceraldehyde-3-phosphate dehydrogenase homo-tetramer formation and mediates its nuclear translocation. Biochim Biophys Acta. 2009;1794(2):254–62.PubMedCrossRefGoogle Scholar
  37. 37.
    Blankschtein ASD. Prediction of critical micelle concentrations and synergism of binary surfactant mixtures containing zwitterionic surfactants. Langmuir. 1997;13(15):3968–81.CrossRefGoogle Scholar
  38. 38.
    Edwards D, Luthy R, Liu Z. Solubilization of polycyclic aromatic hydrocarbons in micellar nonionic surfactant solutions. Environ Sci Technol. 1991;25(1):127–33.CrossRefGoogle Scholar
  39. 39.
    Hait SK, Moulik SP. Determination of critical micelle concentration (CMC) of nonionic surfactants by donor-acceptor interaction with iodine and correlation of CMC with hydrophile–lipophile balance and other parameters of the surfactants. J Surfactants Detergents. 2001;4(3):303–9.CrossRefGoogle Scholar
  40. 40.
    Wolgemuth JL, Workman RK, Manne S. Surfactant aggregates at a flat, isotropic hydrophobic surface. Langmuir. 2000;16(7):3077–81.CrossRefGoogle Scholar
  41. 41.
    Sigma-Aldrich, Inc. Detergents and solubilization reagents. Biofiles. 2008;3(3):30–1.Google Scholar

Copyright information

© Controlled Release Society 2013

Authors and Affiliations

  • Byeong Hee Hwang
    • 1
  • Kenneth Y. Tsai
    • 2
  • Samir Mitragotri
    • 1
  1. 1.Department of Chemical EngineeringUniversity of CaliforniaSanta BarbaraUSA
  2. 2.Departments of Dermatology and ImmunologyUniversity of Texas MD Anderson Cancer CenterHoustonUSA

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