Amino Acids

, Volume 46, Issue 1, pp 153–165 | Cite as

Protein renaturation with simultaneous purification by protein folding liquid chromatography: recent developments

  • Lili Wang
  • Xindu GengEmail author
Invited Review


Protein folding liquid chromatography (PFLC) is a powerful tool for protein refolding with simultaneous purification. We review its recent progress in liquid chromatography and molecular biology, primarily involving the validation of PFLC refolding of proteins containing multiple disulphide bonds, the application of mixed-mode chromatography, PFLC in molecular biology. Representative examples are described.


Protein folding liquid chromatography (PFLC) Proteins Inclusion bodies Refolding with simultaneous purification Mixed-mode liquid chromatography 



Protein folding liquid chromatography


Liquid chromatography


Mixed-mode chromatography

E. coli

Escherichia coli

N state

Native state

U state

Unfold state

M state

Intermediates state


Hydrophobic interaction chromatography


Ion exchange chromatography


Size exclusion chromatography


Affinity chromatography


Reverse phase liquid chromatography


Expanded bed chromatography


Simulated moving bed


Perfusion chromatography


The units of simultaneous renaturation and purification of proteins


Artificial molecular chaperone


AMC-ion exchange chromatography


Guanidine hydrochloride






Hydrophobic interaction expanded bed adsorption chromatography


Immobilised nickel affinity chromatography


A single column

2D column

Two-dimensional column


2D with a single column


Weak cation exchange


Weak anion-exchange chromatography


Stationary phase of hydrophobic interaction chromatography


Recombinant human granulocyte colony stimulating factor


Nipah virus


Hepatitis B virus X


Recombinant human interferon-γ


Recombinant human EGF


Recombinant human stem cell factor


Recombinant alpha-fetoprotein


Recombinant human Flt3 ligand


Recombinant human interferon-alpha 2b


Lipoprotein kringle


Recombinant human vascular endothelial growth factor


Recombinant human Notch ligand delta-like 1





Cyt C

Cytochrome C


Molecular orientation of protein


Stoichiometric displacement theory


Polyethylene glycol


Cellulose-binding domain


Peptidyl prolyl cis trans isomerase


Protein disulphide isomerase




LC–mass spectrometry



This work was supported by the Foundation of Key Laboratory of Modern Separation Science in Shaanxi Province (Nos. 2010JS104; 11JS098), the Foundation of Science and Technology in Shaanxi Province (No. 2010K12-01-05), and the foundation of Provincial Key Description of Analytical Chemistry of Shaanxi Subject (ZDXKL00402).

Conflict of interest

The authors declare that they have no conflict of interest.


  1. Akbari N, Khajeh K, Rezaie S, Mirdamadi S, Shavandi M, Ghaemi N (2010) High-level expression of lipase in Escherichia coli and recovery of active recombinant enzyme through in vitro refolding. Protein Expr Purif 70(1):75–80PubMedCrossRefGoogle Scholar
  2. Alessandra S, Luisa U, Elena M, Sandro P, Alejandro H (2009) Expression and purification of the recombinant mustard trypsin inhibitor 2 (MTI2) in Escherichia coli. J Biosci Bioeng 108(4):282–285CrossRefGoogle Scholar
  3. Anindya B, Leong SSJ (2012) High productivity chromatography refolding process for hepatitis B virus X (HBx) protein guided by statistical design of experiment studies. J Chromatogr A 1223:64–71CrossRefGoogle Scholar
  4. Antonio-Perez A, Ortega-Lopez J (2010) GroEL apical domain, DsbA and DsbC immobilized in cellulose assisted the chromatographic oxidative refolding of lysozyme. J Biotechnol 150S:S85–S86CrossRefGoogle Scholar
  5. Bai Q, Geng XD (2011) Protein folding liquid chromatography, methods in molecular biology, heterologous gene expression in E. coli, vol 705. Humana Press, USA, pp 69–85Google Scholar
  6. Bai Q, Wei YM, Geng MH, Geng XD (1997) Studies on the various refolded intermediates of alpha-amylase denatured by urea with high performance hydrophobic interaction chromatography. Chem J Chin Univ 18:1291–1295Google Scholar
  7. Baneyx F, Mujacic M (2004) Recombinant protein folding and misfolding in Escherichia coli. Nat Biotechnol 22(11):1399–1408PubMedCrossRefGoogle Scholar
  8. Bertoietti L, Regazzoni L, Aldini G, Colomba R, Abballef F, Caccialanza G, Lorenzi ED (2013) Separation and characterisation of beta2-microglobulin folding conformers by ion-exchange liquid chromatography and ion-exchange liquid chromatography–mass spectrometry. Anal Chim Acta 771:108–114CrossRefGoogle Scholar
  9. Bi J, Bai Q, Wang J, Wang LL (2010) Refolding of reduced/ denatured RNase A hydrophobic liquid-solid interface. Chin J Chromatogr 28(8):786–789CrossRefGoogle Scholar
  10. Bradley BS, Atoosa R, Gary SS, Lars K (2011) Temporal development of protein structure during S100A11 folding and dimerization probed by oxidative labeling and mass spectrometry. J Mol Biol 409:669–679CrossRefGoogle Scholar
  11. Cabrita LD, Bottomley SP (2004) Protein expression and refolding–a practical guide to getting the most out of inclusion bodies. Biotechnol Annu Rev 10:31–50PubMedCrossRefGoogle Scholar
  12. Carl F (2007) Protein aggregation processes: in search of the mechanism. Protein Sci 16:2334–2344CrossRefGoogle Scholar
  13. Chen Y, Leong SSJ (2009) Adsorptive refolding of a highly disulfide-bonded inclusion body protein using anion-exchange chromatography. J Chromatogr A 1216:4877–4886PubMedCrossRefGoogle Scholar
  14. Chen Y, Leong SSJ (2010) High productivity refolding of an inclusion body protein using pulsed-fed size exclusion chromatography. Process Biochem 45:1570–1576CrossRefGoogle Scholar
  15. Cherish BPV, Srinivas VK, Krishna MV, Krishna E (2008) Renaturation, purification and characterization of streptokinase expressed as inclusion body in recombinant E. coli. J Chromatogr B 861:218–226CrossRefGoogle Scholar
  16. Choi WC, Kim MY, Suh CW, Lee EK (2005) Solid-phase refolding of inclusion body protein in a packed and expanded bed adsorption chromatography. Process Biochem 40:1967–1972CrossRefGoogle Scholar
  17. Chong FC, Tan WS, Biak DRA, Ling TC, Tey BT (2010) Direct recovery of recombinant nucleocapsid protein of Nipah virus from unclarified Escherichia coli homogenate using hydrophobic interaction expanded bed adsorption chromatography. J Chromatogr A 1217:1293–1297PubMedCrossRefGoogle Scholar
  18. Dasari VKR, Are D, Joginapally VR, Mangamoori LN, Adibhatla KSBR (2008) Optimization of the downstream process for high recovery of rhG-CSF from inclusion bodies expressed in Escherichia coli. Process Biochem 43(5):566–575CrossRefGoogle Scholar
  19. Gao D, Lin DQ, Yao SJ (2008) Patch controlled protein adsorption in mixed-mode chromatography with benzylamine as functional ligand. Biochem Eng J 38:355–361CrossRefGoogle Scholar
  20. Gao D, Tan FC, Wang WP, Wang LL (2013) Resolution enhancement in hydrophobic interaction chromatography via electrostatic interactions. Chin Chem Lett 24:419–421CrossRefGoogle Scholar
  21. Gautam S, Dubey P, Singh P, Kesavardhana S, Varadarajan R, Gupta MN (2012) Smart polymer mediated purification and recovery of active proteins from inclusion bodies. J Chromatogr A 1235:10–25PubMedCrossRefGoogle Scholar
  22. Geng XD, Wang CZ (2007) Protein folding liquid chromatography and its recent developments. J Chromatogr B 849:69–80CrossRefGoogle Scholar
  23. Geng XD, Wang LL (2008) Liquid chromatography of recombinant proteins and protein drugs. J Chromatogr B 866:133–153CrossRefGoogle Scholar
  24. Geng XD, Zhang YJ (2008b) A kind of chromatographic cake and its manufacturing method and application, EP 1 396 721 B1Google Scholar
  25. Geng XD, Guo LA, Chang JH (1990) Study of the retention mechanism of proteins in hydrophobic interaction chromatography. J Chromatogr A 507:1–23CrossRefGoogle Scholar
  26. Geng XD, Bai Q, Wang CZ (2006) Protein folding liquid chromatography. Sci Press, Beijing, pp 175–177Google Scholar
  27. Geng XD, Ke CY, Chen G, Liu P, Wang F, Zhang HQ, Sun X (2009) On-line separation of native proteins by two-dimensional liquid chromatography using a single column. J Chromatogr A 1216:3553–3562PubMedCrossRefGoogle Scholar
  28. Gorka J, Rohmer M, Bornemann S, Papasotiriou DG, Baeumlisberger D, Arrey TN, Bahr U, Karas M (2012) Perfusion reversed-phase high-performance liquid chromatography for protein separation from detergent-containing solutions: an alternative to gel-based approaches perfusion chromatography. Anal Biochem 424:97–107PubMedCrossRefGoogle Scholar
  29. Gueorguieva L, Palani S, Rinas U, Jayaraman G, Seidel-Morgenstern A (2011) Recombinant protein purification using gradient assisted simulated moving bed hydrophobic interaction chromatography. Part II: process design and experimental validation. J Chromatogra A 1218:6402–6411CrossRefGoogle Scholar
  30. Hil’chuk PV, Okuniev OV, Pavlova MV, Irodov DM, Horbatiuk OB (2006) Obtaining of ScFv-CBD fusion protein and its application for affinity purification of recombinant human interferon alpha-2b. Ukr Biokhim Zh 78(2):52–61PubMedGoogle Scholar
  31. Hou Y, Hansen TB, Staby A, Cramera SM (2010) Effects of urea induced protein conformational changes on ion exchange chromatographic behavior. J Chromatogr A 1217:7393–7400PubMedCrossRefGoogle Scholar
  32. Ito L, Okumura M, Tao K, Kasai Y, Tomita S, Oosuka A, Yamada H, Shibano T, Shiraki K, Kumasaka T, Yamaguchi H (2012) Glutathione ethylester, a novel protein refolding reagent, enhances both the efficiency of refolding and correct disulfide formation. Protein J 31:499–503PubMedCrossRefGoogle Scholar
  33. Jia J (2010) Refolding and simultaneous purification of the recombinant human Flt3 ligand by protein folding liquid chromatography. Masteral degree thesis, Northwest University, Xi’an, pp 49–54Google Scholar
  34. Jia J, Wang LL, Gao D, Geng XD (2010) Refolding and simultaneous purification of the recombinant human Flt3 ligand from inclusion bodies by high performance hydrophobic interaction chromatography. Chin J Chromatogr 28:535–540CrossRefGoogle Scholar
  35. Jia X, Ke C, Geng XD (2012) Studies on the characterization of stationary phase for protein retention on mixed-mode chromatography. Acta Chim Sinica 70:1631–1636CrossRefGoogle Scholar
  36. Ke CY, Meng ZC (2012) Denatured ribonuclease refolding by glutathione bonding column. Chem J Chin Univ 33:925–930Google Scholar
  37. Ke CY, Li JJ, Liu ZL, Geng XD (2009) A new approach for characterizing the intermediate feature of α-chymotrypsin refolding by hydrophobic interaction chromatography. Int J Mol Sci 10:616–628PubMedCentralPubMedCrossRefGoogle Scholar
  38. Ke CY, Sun WJ, Zhang QZ, Geng XD (2013) Refolding of urea-denatured α-chymotrypsin by protein-folding liquid chromatography. Biomed Chromatogr 27:433–439CrossRefGoogle Scholar
  39. Kennedy LA, Kopaciewicz W, Regnier FE (1986) Multimodal liquid chromatography columns for the separation of proteins in either the anion-exchange or hydrophobic-interaction mode. J Chromatogr 359:73–84PubMedCrossRefGoogle Scholar
  40. Kimberly AK, Charles HS, Shelly AP (2010) Industrial case study: evaluation of a mixed-mode resin for selective capture of a human growth factor recombinantly expressed in E. coli. J Chromatogr A 1217:235–242CrossRefGoogle Scholar
  41. Li X, Leong SSJ (2011) A chromatography-focused bioprocess that eliminates soluble aggregation for bioactive production of a new antimicrobial peptide candidate. J Chromatogr A 1218:3654–3659PubMedCrossRefGoogle Scholar
  42. Li JJ, Wang AQ, Janson JC, Ballagi A, Chen J, Liu YD, Ma GH, Su ZG (2009) Immobilized Triton X-100-assisted refolding of green fluorescent protein-tobacco etch virus protease fusion protein using β-cyclodextrin as the eluent. Process Biochem 44:277–282CrossRefGoogle Scholar
  43. Li M, Fan H, Liu J, Wang M, Wang LL, Wang CZ (2012a) High pH solubilization and chromatography-based renaturation and purification of recombinant human granulocyte colony-stimulating factor from inclusion bodies. Appl Biochem Biotechnol 166(5):1264–1274PubMedCrossRefGoogle Scholar
  44. Li J, Zheng L, Li PL, Wang FS (2012b) Intein-mediated expression, purification, and characterization of thymosin α1-thymopentin fusion peptide in Escherichia coli. Protein Expr Purif 84:1–8PubMedCrossRefGoogle Scholar
  45. Lin S, Karger BL (1990) Reversed-phase chromatographic behavior of proteins in different unfolded states. J. Chromatogr 499:89–102PubMedCrossRefGoogle Scholar
  46. Liu XQ, Yang XQ, Xie FH, Song LY, Zhang GQ, Qian SJ (2007) On-column refolding and purification of transglutaminase from Streptomyces fradiae expressed as inclusion bodies in Escherichia coli. Protein Expr Purif 51:179–186PubMedCrossRefGoogle Scholar
  47. Liu P, Yang HY, Geng XD (2009a) Mixed retention mechanism of proteins in weak anion-exchange chromatography. J Chromatogr A 1216:7497–7504PubMedCrossRefGoogle Scholar
  48. Liu ZL, Ke CY, Geng XD (2009b) Influence of high-performance hydrophobic interaction chromatographic stationary phase on refolding of urea-denatured α-chymotrypsin. Chin J Anal Chem 28:34–37Google Scholar
  49. Lu BY, Chang JY (2010) Rapid and irreversible reduction of protein disulfide bonds. Anal Biochem 405:67–72PubMedCrossRefGoogle Scholar
  50. Lu ML, Cao X, Yang X, Zheng H, Liu N, Jiang Y, Lin DH, Chen YJ (2010) A diketoreductase exhibits unique renaturation profile from thermal-induced protein unfolding. Amino Acids 39:609–613PubMedCrossRefGoogle Scholar
  51. Lu HL, Lin DQ, Gao D, Yao SJ (2013) Evaluation of immunoglobulin adsorption on the hydrophobic charge-induction resins with different ligand densities and pore sizes. J Chromatogr A 1278:61–68PubMedCrossRefGoogle Scholar
  52. Luo M, Guan YX, Yao SJ (2011) On-column refolding of denatured lysozyme by the conjoint chromatography composed of SEC and immobilized recombinant DsbA. J Chromatogr B 879:2971–2977CrossRefGoogle Scholar
  53. Majtan T, Kraus JP (2012) Folding and activity of mutant cystathionine b-synthase depends on the position and nature of the purification tag: Characterization of the R266K CBS mutant. Protein Expr Purif 82:317–324PubMedCentralPubMedCrossRefGoogle Scholar
  54. Mehander WR, Rassi ZE, Horvath C (1989) Interplay of hydrophobic and electrostatic interactions in biopolymer chromatography: effect of salts on the retention of proteins. J Chromatogr A 469:3–27CrossRefGoogle Scholar
  55. Metcalf D (1988) Colony stimulating factors and hemopoiesis. Ann Acad Med Singap 17(2):166–170PubMedGoogle Scholar
  56. Murugan E, Kong R, Sun HH, Rao F, Liang ZX (2010) Expression, purification and characterization of the acyl carrier protein phosphodiesterase from Pseudomonas aeruginosa. Protein Expr Purif 71:132–138PubMedCrossRefGoogle Scholar
  57. Nageswara RP, Gullipalli D, Abani KB (2009) Bacterially expressed recombinant WD40 domain of human Apaf-1. Protein Expr Purif 67:53–60CrossRefGoogle Scholar
  58. Park BJ, Lee CH, Mun SY, Koo YM (2006) Novel application of simulated moving bed chromatography to protein refolding. Process Biochem 41:1072–1082CrossRefGoogle Scholar
  59. Petrov S, Nacheva G, Ivanov I (2010) Purification and refolding of recombinant human interferon-gamma in urea–ammonium chloride solution. Protein Expr Purif 73:70–73PubMedCrossRefGoogle Scholar
  60. Phadtare S, Fisher MT, Yarbrough LR (1994) Refolding and release of tubulins by a functional immobilized GroEL column. Biochimica et Biophysica Acta 1208:189–192PubMedCrossRefGoogle Scholar
  61. Radford SE, Dobson CM, Evans PA (1992) The folding of hen lysozyme involves partially structured intermediates and multiple pathways. Nature 358:302–307PubMedCrossRefGoogle Scholar
  62. Raghava S, Barua B, Singh PK, Das M, Madan L, Bhattacharyya S, Bajaj K, Gopal B, Varadarajan R, Gupta MN (2008) Refolding and simultaneous purification by three-phase partitioning of recombinant proteins from inclusion bodies. Protein Sci 17(11):1987–1997PubMedCrossRefGoogle Scholar
  63. Rinas U, Hoffmann F, Betiku E, Estapé D, Marten S (2007) Inclusion body anatomy and functioning of chaperone-mediated in vivo inclusion body disassembly during high-level recombinant protein production in Escherichia coli. J Biotechnol 127:244–257PubMedCrossRefGoogle Scholar
  64. Robert SC, Richard RB (2010) Expression, purification, and refolding of active Nrf2 transcription factor fused to protein transduction TAT tag. Protein Expr Purif 74:280–288CrossRefGoogle Scholar
  65. Schlegl R, Iberer G, Machold C, Necina R, Jungbauer A (2003) Continuous matrix-assisted refolding of proteins. J Chromatogr A 1009:119–132PubMedCrossRefGoogle Scholar
  66. Sharapova OA, Yurkova MS, Laurinavichyute DK, Andronova SM, Fedorov AN, Severin SE, Severin ES (2011) Efficient refolding of a hydrophobic protein with multiple S–S bonds by on-resin immobilized metal affinity chromatography. J Chromatogr A 1218:5115–5119PubMedCrossRefGoogle Scholar
  67. Sharma K, Cherish Babu PV, Sasidhar P, Srinivas VK, Mohan VK, Krishna E (2008) Recombinant human epidermal growth factor inclusion body solubilization and refolding at large scale using expanded-bed adsorption chromatography from Escherichia coli. Protein Expr Purif 60:7–14PubMedCrossRefGoogle Scholar
  68. Shi ZX, He F, Wang LL, Liang YM, Han H, Wang CZ, Zhao Q, Geng XD (2008) Expression, refolding, and purification of a truncated human delta-like 1, a ligand of Notch receptors. Protein Expr Purif 59:242–248PubMedCrossRefGoogle Scholar
  69. Špela P, Radovan K (2011) Active protein aggregates produced in Escherichia coli. Int J Mol Sci 12:8275–8287CrossRefGoogle Scholar
  70. Sun X, Yang Y, Geng XD (2010) Effect of hydrophobic interaction on protein retention in weak-cation exchange chromatography. Chin J Anal Sci 26:6–10Google Scholar
  71. Sun WM, Dai XX, Zheng YP, Wang JW, Hou LL, Du J, Hua HG (2011) On-column refolding purification of DT389-hIL13 recombinant protein expressed in Escherichia coli. Protein Expr Purif 75:83–88PubMedCrossRefGoogle Scholar
  72. Swaminathan R, Ravi VK, Kumar S, Kumar MVS, Chandra N (2011) Lysozyme: a model protein for amyloid research. Adv Protein Chem Struct Biol 84:63–111PubMedCrossRefGoogle Scholar
  73. Swietnicki W (2006) Folding aggregated proteins into functionally active forms. Curr Opin Biotechnol 17:367–372PubMedCrossRefGoogle Scholar
  74. Tong HF, Lin DQ, Gao D, Yuan XM, Yao SJ (2013) Caprylate as the albumin-selective modifier to improve IgG purification with hydrophobic charge-induction chromatography. J Chromatogr 1285:88–96CrossRefGoogle Scholar
  75. Valko KV, Snyder LR, Glajch JL (1993) Retention in reversed-phase liquid chromatography as a function of mobile-phase composition. J Chromatogr A 656:501–520CrossRefGoogle Scholar
  76. Vermasvuori R, Koskinen J, Salonen K, Sirén N, Weegar J, Dahlbacka J, Kalkkinen N, von Weymarn N (2009) Production of recombinant HIV-1 nef protein using different expression host systems: A techno-economical comparison. Biotechnol Prog 25:95–102PubMedCrossRefGoogle Scholar
  77. Walsh G (2010) Biopharmaceutical benchmarks. Nat Biotechnol 28:917–924PubMedCrossRefGoogle Scholar
  78. Wang CZ, Geng XD (2012) Refolding and purification of recombinant human granulocyte colony-stimulating factor using hydrophobic interaction chromatography at a large scale. Process Biochem 47:2262–2266CrossRefGoogle Scholar
  79. Wang DM, Shi LM (2009) High-level expression, purification, and in vitro refolding of soluble tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). Appl Biochem Biotechnol 157:1–9PubMedCrossRefGoogle Scholar
  80. Wang CZ, Wang LL, Geng XD (2006) Renaturation of recombinant human granulocyte colony-stimulating factor produced from Escherichia coli using size exclusion chromatography. J Liq Chromatogr Relat Technol 29:203–217CrossRefGoogle Scholar
  81. Wang CZ, Wang LL, Geng XD (2007) Renaturation with simultaneous purification of rhG-CSF from Escherichia coli by ion exchange chromatography. Biomed Chromatogr 21:1291–1296PubMedCrossRefGoogle Scholar
  82. Wang CZ, Wang LL, Geng XD (2008a) High recovery refolding of rhG-CSF from Escherichia coli using urea gradient size exclusion chromatography. Biotechnol Prog 24:209–213PubMedCrossRefGoogle Scholar
  83. Wang CZ, Liu JH, Wang LL, Geng XD (2008b) Solubilization and refolding with simultaneous purification of recombinant human stem cell factor. Appl Biochem Biotechnol 144:181–189PubMedCrossRefGoogle Scholar
  84. Wang HJ, Dai JX, Li BH, Fan KX, Peng L, Zhang DP, Cao ZG, Qian WZ, Wang H, Zhao J, Guo YJ (2008c) Expression, purification, and characterization of an immunotoxin containing a humanized anti-CD25 single-chain fragment variable antibody fused to a modified truncated Pseudomonas exotoxin A. Protein Expr Purif 58:140–147PubMedCrossRefGoogle Scholar
  85. Wang CZ, Wang LL, Geng XD (2009a) Optimization of refolding with simultaneous purification of recombinant human granulocyte colony-stimulating factor from Escherichia coli by immobilized metal ion affinity chromatography. Biochem Eng J 43:197–202CrossRefGoogle Scholar
  86. Wang FW, Liu YD, Ma GH, Su ZG (2009b) Glycerol-assisted hydrophobic interaction chromatography improving refolding of recombinant human granulocyte colony-stimulating factor. Appl Biochem Biotechnol 159:634–641PubMedCrossRefGoogle Scholar
  87. Wang CZ, Zhang QM, Cheng Y, Wang LL (2010a) Refolding of denatured/reduced lysozyme at high concentrations by artificial molecular chaperone-ion exchange chromatography. Biotechnol Prog 26(4):1073–1079PubMedGoogle Scholar
  88. Wang H, Liu XX, He YS, Dong JX, Sun YM, Liang Y, Yang JY, Lei HT, Shen YD, Xu XY (2010b) Expression and purification of an anti-clenbuterol single chain Fv antibody in Escherichia coli. Protein Expr Purif 72:26–31PubMedCrossRefGoogle Scholar
  89. Wang GZ, Dong XY, Sun Y (2011a) Ion-exchange resins greatly facilitate refolding of like-charged proteins at high concentrations. Biotechnol Bioeng 105:1068–1077CrossRefGoogle Scholar
  90. Wang LL, Wang CZ, Geng XD (2011b) Fast preparation of recombinant human stem cell factor from inclusion bodies using high performance hydrophobic interaction chromatography. Chin J Chromatogr 29:36–41CrossRefGoogle Scholar
  91. Wang F, Min Y, Geng XD (2012) Fast separations of intact proteins by liquid chromatography. J Sep Sci 35:3033–3045PubMedCrossRefGoogle Scholar
  92. Waugh DS (2005) Making the most of affinity tags. Trends Biotechnol 23:316–320PubMedCrossRefGoogle Scholar
  93. Wu X (2013) Notch ligand delta-like1 fusion protein expression, and refolding with simultaneous purification. Master’s thesis. College of chemistry and materials science, Northwest University, Xi’an, pp 38–46Google Scholar
  94. Wu D, Wang CZ, Geng XD (2007) An approach for increasing the mass recovery of proteins derived from inclusion bodies in biotechnology. Biotechnol Prog 23:407–413PubMedCrossRefGoogle Scholar
  95. Wu D, Gao D, Bai Q, Geng XD (2008) Preparation and identification of recombinant human interferon-γ. Chin J Chromatogr 26(2):206–211Google Scholar
  96. Yang Y, Geng XD (2011) Mixed-mode chromatography and its applications to biopolymers. J Chromatogr A 1218:8813–8825PubMedCrossRefGoogle Scholar
  97. Yang X, Yu W, Geng XD (2013) On-line fast purification of cytochrome C from bovine pancreas by two-dimensional liquid chromatography using a single column. Sci China (Ser B) Chin 43(5):599–609Google Scholar
  98. Zhang YH, Ma YS, Yang MY, Min SJ, Yao JM, Zhu LJ (2011) Expression, purification, and refolding of a recombinant human bone morphogenetic protein-2 in vitro. Protein Expr Purif 75:155–160PubMedCrossRefGoogle Scholar
  99. Zhou K (2009) Synthesis of new guanidine protein folding chromatographic stationary phase and its effect on the refolding and purification of rhG-CSF. Master’s thesis. College of chemistry and materials science, Northwest University, Xi’an, pp 31–43Google Scholar

Copyright information

© Springer-Verlag Wien 2013

Authors and Affiliations

  1. 1.Key Laboratory of Synthetic and Natural Functional Molecular Chemistry of Ministry of Education, Shaanxi Key Laboratory of Modern Separation Science, Institute of Modern Separation ScienceNorthwest UniversityXi’anChina

Personalised recommendations