Biocomplex purification

Biocomplex provides purification technologies and expertise specializing in preparative ultracentrifugation, monolithic chromatography and asymmetrical flow field-flow fractionation (AF4). Provided techniques can be used in different combinations to yield high quality samples in quantity. Biosafety level 1 and 2 organisms can be processed.

• Ultracentrifugation combined with swing-out and fixed-angle rotors of different capacities;
• Asymmetrical flow field-flow fractionation (AF4);
• Monolithic ion exchange chromatography;
• User training, assisted use of instruments, open access for trained users, specialist support and maintenance;
• Project planning;
• Full service packages on sample purification (including analysis of the quality and quantity);
• Collaborative projects;
• Courses in biomolecular complex purification

Ultracentrifugation service

The preparative ultracentrifugation service provides facilities for rate-zonal, equilibrium, flotation and differential centrifugation. It is equipped with a farm of ultracentrifuges as well as swing out and fixed angle rotors with different capacities. We also have instrumentation for gradient making and provide fractionation services. Superspeed centrifuges are available also for the users.

Asymmetrical flow field-flow fractionation (AF4) service

AF4 provides size-based separation of sample components. It is applicable for separation of macromolecules, colloids as well as nano- and microparticles. Separation is done in liquid phase without stationary phase. The benefits of AF4 are the following:

• Gentle size-based separation in liquid phase;
• Large dynamic size range;
• Rapid analysis time;
• High yields

Chromatography service

CIM® monolithic columns can be utilized for purification of viruses, virus-like particles, nucleic acids, proteins and their complexes. We routinely have weak (DEAE) and strong (QA) anion exchange columns. Please ask for other column types. The benefits of CIM columns are the following:

• Large pore sizes enable entering and binding of large macromolecular complexes to the matrix;
• High flow rates enable fast separation;
• High yields and sample concentrations can be achieved.

Please find the downloadable poster of Bíocomplex services below.

• High quality purification of large and fragile macromolecular complexes in quantity;
• Purification based on differences in sedimentation rate (mass, size, shape)  or density (ultracentrifugation), shape or size (AF4), and surface charge (monoliths);
• Separation and purification of cells, organelles, macromolecules, macromolecular complexes, bacteria, vesicles and viruses etc.

Ultracentrifuges

• Five ultracentrifuges (Thermo Scientific Sorvall WX Ultra Series);
• Swing-out and fixed angle rotors with different capacities (Sorvall and Beckman), see table.
• Instrumentation for gradient making (IBS INTEGRA Biosciences peristaltic pump; BioComp Gradient master)
• Gradient fractionator (BioComp Piston Gradient Fractionator, only provided as a service)
• Beckmann Airfuge for fast pelleting of small sample volumes.
  • A-95 fixed angle rotor: 4*450 µl
  • A-110 fixed angle rotor: 6*175 µl

Super-speed centrifugation (Thermo Scientific)

• Sorvall LYNX4000
• Superspeed rotors
  • Fiberlite F20-12x50 LEX, 13 000 rpm
  • Fiberlite F12-6x500 LEX, 8000 rpm 
  • Fiberlite F14-6x250y, 10 000 rpm

AF2000 MultiFlow (Postnova)

• Analytical (CHA-612) and semi-preparative (CHA-601) separation channel (semi-preparative available in 2019; temperature controlled +5 - +75 °C)
• Regenerated cellulose membranes (MWCOs 1, 10 and 100 kDa)
• Spacers (thickness 250, 350, and 500 µm)
• UV-Vis Detector (PN3211; wavelength 190-600 nm)
• Smart stream splitter (PN1650)
• Fraction collector (PN8050; temperature controlled +10 - +40 °C)
• Autosampler (available in 2019)

CIM monolithic ion-exchange columns

• DEAE and QA anion exchange columns (BiaSeparations)
• Lab-scale chromatography system (New ÄKTAPure 25M installed in October 2019)

• Our service is open for all users;
• Our facility can be used for organisms of the biosafety levels 1 and 2;
• We serve on a first-come-first-served basis. Access is prioritized based on the scientific feasibility peer-review of the project summary within the units;
• New from the beginning of 2019: Register your project by filling the Biocomplex User Registration e-form https://elomake.helsinki.fi/lomakkeet/93251/lomake.html for approval (feasibility, technical checking; safety issues) and contact grp-biocomplexservice[at]helsinki.fi. Note that the registrations are valid for one calendar year;
• We recommend users to register as a member of Instruct-ERIC (Instruct registration);
• Training on the safety and operation issues is obligatory before the first use. After your project has been approved by the Biocomplex personnel, please contact us for reserving the time for training (grp-biocomplexservice[at]helsinki.fi);
• Users are expected to clean up the work places immediately after they have completed their work; 

• Acknowledgements: User agrees to acknowledge research infrastructures (RIs) in the upcoming publications and theses with i.e. the following text: “The use of the facilities and expertise of the Instruct-HiLIFE Biocomplex unit, member of Biocenter Finland and Instruct-FI, is gratefully acknowledged." In addition, users agree to send the bibliography of the published material (scientific papers, theses) to biocomplexservice[at]helsinki.fi.

  biocomplex_project_registration_uh.docx

Biocomplex services are no longer free of charge. Fees will be collected to cover some of the maintenance costs of the Biocomplex Unit. The prices are available below. Note that the prices do not include VAT.

Centrifugation

• Do you borrow the rotors for centrifuges?

For safety reasons we do not borrow our rotors.

• I have obtained the training for centrifuges from another unit/university. Do I need the training from Biocomplex unit?

Yes you do. This for your own safety and for the safety of our customers and other personnel.

• I am an experienced centrifuge user. Can I do the training?

Unfortunately not. This for your own safety and for the safety of our customers and other personnel.

Asymmetrical flow field-flow fractionation

• How many samples can be analyzed by the same membrane used in AF4?

Obviously this depends on the sample that is being analyzed, but the manufacturer's estimate is 30-50 analysis per one membrane.

• How many AF4 analysis can be performed during one working day?

This depends on the type of analysis that is done. Minimum number is ~7 analysis, if the anaysis time is ~1 hour.

Monolithic ion exchange chromatography

• Do you provide other column types as well?

Currently, we only have the DEAE and QA chemistries available routinely. If you have other types of columns in mind, please contact us.

User policies

• Does the PI of the project need to fill in his/her own Biocomplex service request form?

No. It is enough that the PI signs the service request form that is filled in by the User. By signing the PI agrees to cover the costs from the service.

• I have already registered as a Biocomplex user for 2018. Do I need to register again for 2019?

Yes you do. From 2019 we start to collect the registration forms electronically. While renewing your registration, please check that the information for invoicing is still valid?

Publications with acknowledgements to the Biocomplex Unit (Instruct Centre for Virus and Macromolecular Complex Production, ICVIR 2009-2017)

2019

2018

• Adriaenssens EM, Wittmann J, Kuhn JH, Turner D, Sullivan MB, Dutilh BE, Jang HB, van Zyl LJ, Klumpp J, Lobocka M, Moreno Switt AI, Rumnieks J, Edwards RA, Uchiyama J, Alfenas-Zerbini P, Petty NK, Kropinski AM, Barylski J, Gillis A, Clokie MRC, Prangishvili D, Lavigne R, Aziz RK, Duffy S, Krupovic M, Poranen MM, Knezevic P, Enault F, Tong Y, Oksanen HM, Rodney Brister J. 2018. Taxonomy of prokaryotic viruses: 2017 update from the ICTV Bacterial and Archaeal Viruses Subcommittee. Arch Virol. 163:1125-1129.
• Atanasova NS, Demina TA, Krishnam Rajan Shanthi SV, Oksanen HM, and Bamford DH. 2018. Extremely halophilic pleomorphic archaeal virus HRPV9 extends the diversity of pleolipoviruses with integrases. Res Microbiol. 169:500-504 doi: 10.1016/j.resmic.2018.04.004.
• Atanasova NS, Heiniö CHE, Demina T, Bamford DH, Oksanen HM. 2018. The unexplored diversity of pleolipoviruses: the surprising case of two viruses with identical major structural modules. Genes 9:131.
• Azinas S, Bano F, Torca I, Bamford DH, Schwart GA, Esnaola J, Oksanen HM, Richter R, Abrescia NGA. 2018. Membrane-containing virus particle exhibits mechanics of a composite material for genome protection. Nanoscale 10(16):7769-7779.
• Duyvesteyn HME, Ginn HM, Pietilä MK, Wagner A, Hattne J, Grimes JM, Hirvonen E, Evans G, Parsy M-L, Sauter NK, Brewster AS, Huiskonen JT, Stuart DI, Sutton G, Bamford DH. 2018. Towards in cellulo virus crystallography. Sci. Rep. 8:3771.
• Eskelin K, Poranen MM. 2018. Controlled disassembly and purification of functional viral subassemblies using asymmetrical flow field-flow fractionation (AF4). Viruses, 10(11), 579; https://doi.org/10.3390/v10110579
• Lampi M, Oksanen HM, Meier F, Moldenhauer E, Poranen MM, Bamford DH, and Eskelin K. 2018. Asymmetrical flow field-flow fractionation in purification of an enveloped bacteriophage ɸ6. J Chrom B. 1095: 251-257.
• Leigh B, Breitbart M, Oksanen HM, Bamford DH & Dishaw L. 2018. Genome sequence of PM2-like phage Cr39582 induced from Pseudoalteromonas sp. isolated from the gut of Ciona robusta. Genome Announc 6(21): e00368-18.
• Levanova A, Poranen MM. 2018. Application of steric exclusion chromatography on monoliths for separation and purification of RNA molecules. J Chrom A.1574:50-59. doi: 10.1016/j.chroma.2018.08.063.
• Luhtanen AM, Eronen-Rasimus E, Oksanen HM, Tison J-L, Delille B, Dieckmann G, Rintala J-M, Bamford DH. 2018. The first known virus isolates from Antarctic sea ice have complex infection patterns. FEMS Microbiol Ecol. 94:fiy028. 
• Niehl A, Soininen M, Poranen MM, Heinlein M. 2018. Synthetic biology approach for plant protection using dsRNA. Plant Biotechnol J. 16(9): 1679-1687. doi: 10.1111/pbi.12904
• Pietilä MK, van Hemert MJ, Ahola T. 2018. Purification of highly active alphavirus replication complexes demonstrates altered fractionation of multiple cellular membranes. J Virol. 92:e01852-17.
• Sun X, Ilca SL, Huiskonen JT, Poranen MM. 2018. Dual role of a viral polymerase in viral genome replication and particle self-assembly. mBio. 9(5): e01242-18. doi: 10.1128/mBio.01242-18.

2017

• Adriaenssens EM, Krupovic M, Knezevic P, Ackermann HW, Barylski J, Brister JR, Clokie MR, Duffy S, Dutilh BE, Edwards RA, Enault F, Jang HB, Klumpp J, Kropinski AM, Lavigne R, Poranen MM, Prangishvili D, Rumnieks J, Sullivan MB, Wittmann J, Oksanen HM, Gillis A, Kuhn JH. 2017. Taxonomy of prokaryotic viruses: 2016 update from the ICTV bacterial and archaeal viruses subcommittee. Arch. Virol. 162:1153-1157.
• Coloma SE, Dienstbier A, Bamford DH, Sivonen K, Roine E, Hiltunen T. 2017. Newly isolated Nodularia phage influences cyanobacterial community dynamics. Environ. Microbiol. 19:273-286.
• Demina TA, Pietilä MK, Svirskaitė J, Ravantti JJ, Atanasova NS, Bamford DH, Oksanen HM. 2017. HCIV-1 and other tailless icosahedral internal membrane-containing viruses of the family Sphaerolipoviridae. Viruses. 9:32.
• Eskelin K., Lampi M, Meier F, Moldenhauer E, Bamford DH, and Oksanen HM. Halophilic viruses with varying biochemical and biophysical properties are amenable to purification with asymmetrical flow field-flow fractionation. Extremophiles. 21:1119-1132.
• Hanhijärvi KJ, Ziedaite G, Bamford DH, Hæggström E, Poranen MM. 2017. Single-molecule measurements of viral ssRNA packaging. RNA. 23:119-129.
• Pietilä MK, Albulescu IC, Hemert MJV, Ahola T. 2017. Polyprotein processing as a determinant for in vitro activity of Semliki Forest virus replicase. Viruses. 9:292.
• Prangishvili D, Bamford DH, Forterre P, Iranzo J, Koonin EV, Krupovic M. 2017. The enigmatic archaeal virosphere. Nat. Rev. Microbiol. 15:724-739.
• Sinclair RM, Ravantti JJ, Bamford DH. 2017. Nucleic and amino acid sequences support structure-based viral classification. J. Virol. 91:e02275-16.                                    
• Santos-Pérez I, Oksanen HM, Bamford DH, Goñi FM, Reguera D, Abrescia NG. 2017. Membrane-assisted viral DNA ejection. Biochim. Biophys. Acta. 1861:664-672.
• Sun Z, El Omari K, Sun X, Ilca SL, Kotecha A, Stuart DI, Poranen MM, Huiskonen JT. 2017. Double-stranded RNA virus outer shell assembly by bona fide domain-swapping. Nat. Commun. 8:14814

2016

• Atanasova NS, Bamford DH, Oksanen HM. 2016. Virus-host interplay in high salt environments. Environmental microbiology reports. Environ. Microbiol. Rep. 8:431-44.
• Demina TA, Atanasova NS, Pietilä MK, Oksanen HM, Bamford DH. 2016. Vesicle-like virion of Haloarcula hispanica pleomorphic virus 3 preserves high infectivity in saturated salt. Virology 499:40-51.
• Demina TA, Pietilä MK, Svirskaite J, Ravantti JJ, Atanasova NS, Bamford DH, Oksanen HM. 2016. Archaeal virus HCIV-1 highlights conserved elements in icosahedral membrane-containing DNA viruses from extreme environments. mBio 7(4):e00699-16.
• Eskelin K, Lampi M, Meier F, Moldenhauer E, Bamford DH, Oksanen HM. 2016. Asymmetric flow field flow fractionation methods for virus purification. J. Chromatogr. A. 1469:108-119.
• Hanhijärvi KJ, Ziedaite G, Hæggström E, Bamford DH. 2016. Temperature and pH dependence of DNA ejection from archaeal lemon-shaped virus His1. Eur. Biophys. J. 45:435-442.
• Ivanov KI, Eskelin K, Bašić M, De S, Lõhmus A, Varjosalo M, Mäkinen K. 2016. Molecular insights into the function of the viral RNA silencing suppressor HCPro. Plant. J. 85:30-45.
• Orentaite, I., Poranen, M. M., Oksanen, H. M., Daugelavicius, R. & Bamford, D. H. 2016. K2 killer toxin-induced physiological changes in the yeast Saccharomyces cerevisiae. FEMS Yeast Research. 16:1-8.  
• Pietilä MK, Roine E, Sencilo A, Bamford DH, Oksanen HM. 2016. Pleolipoviridae, a newly proposed family comprising archaeal pleomorphic viruses with single-stranded or double-stranded DNA genomes. Arch. Virol. 161:249-56.
• Svirskaitė J, Oksanen HM, Daugelavičius R, Bamford DH. 2016. Monitoring Physiological Changes in Haloarchaeal Cell during Virus Release. Viruses. 8(3). pii: E59.

2015

• Atanasova NS, Oksanen HM, and Bamford DH. 2015. Haloviruses of archaea, bacteria, and eukaryotes. Curr. Opin. Microbiol. 25:40-48.
• Atanasova NS, Bamford DH, Oksanen HM. 2015. Haloarchaeal virus morphotypes. Biochimie. 118:333-343.
• Atanasova NS, Demina TA, Buivydas A, Bamford DH, and Oksanen HM. 2015. Archaeal viruses multiply: temporal screening in a solar saltern. Viruses. 7:1902-1926.
• Atanasova NS, Senčilo A, Pietilä MK, Roine E, Oksanen HM, and Bamford DH. 2015. Comparison of lidip-containing bacterial and archaeal viruses. Adv. Virus Res. 92:1-61.
• Gil-Carton D, Jaakkola ST, Charro D, Peralta B, Castaño-Díez, Oksanen HM, Bamford DH, Abrescia NGA. 2015. Insight into the assembly of viruses with vertical single β-barrel major capsid proteins. Structure. 23:1866-1877.
• Hong C, Pietilä MK, Fu C, Schmid MF, Bamford DH, and Chiu W. 2015. Lemon-shaped halo archaeal virus His1 with uniform tail but variable capsid structure. Proc. Natl. Acad. Sci USA 112:2449-2454.
• Karttunen J, Mäntynen S, Ihalainen TO, Bamford JKH and Oksanen HM. 2015. Non-structural proteins P17 and P33 are involved in the assembly of the internal membrane-containing virus PRD1. Virology. 482:225-233.
• Liu Y, Wang J, Liu Y, Wang Y, Zhang Z, Oksanen HM, Bamford DH and Chen X. 2015. Identification and characterization of SNJ2, the first temperate pleolipovirus integrating into the genome of the SNJ1-lysogenic archaeal strain. Mol. Microbiol. 98:1002-1020.
• Mattila SP, Oksanen HM, and Bamford JKH. 2015. Probing of protein interactions in the membrane-containing virus PRD1. J. Gen. Virol. 96: 453-462.
• Mäntynen S, Laanto E, Kohvakka A, Poranen MM, Bamford JK, Ravantti JJ. 2015. New enveloped dsRNA phage from freshwater habitat. J. Gen. Virol. 96:1180-1189.
• Poranen MM, Bamford DH, and Oksanen HM. 2015. Membrane-containing bacteriophages. In: eLS. John Wiley & Sons, Ltd: Chichester. DOI: 10.1002/9780470015902.a0000779.pub3
• Quemin ERJ, Pietilä MK, Oksanen HM, Forterre P, Rijpstra WIC, Schouten S, Bamford DH, Prangishvili D, Krupovic M. 2015. Sulfolobus spindle-shaped virus 1 contains glycosylated capsid proteins, a cellular chromatin protein and host-derived lipids. J Virol. 15:11681-11691.
• Sencilo A, Luhtanen A-M, Saarijärvi M, Bamford DH, Roine E. 2015. Cold-active bacteriophages from the Baltic Sea ice have diverse genomes and virus-host interactions. Environ Microbiol. 17:3628-41.

2014

• Hong C, Oksanen HM, Liu X, Jakana J, Bamford DH, and Chiu W. 2014. A structural model of the genome packaging process in a membrane-containing double stranded DNA virus. PLoS Biol. 12:e1002024.
• Krupovic M, Bamford DH, Koonin EV. 2014. Conservation of major and minor jelly-roll capsid proteins in Polinton (Maverick) transposons suggests that they are bona fide viruses. Biol. Direct. 9:6
• Luhtanen A-M, Eronen-Rasimus E, Kaartokallio H, Rintala J-M, Autio R, and Roine E. 2014. Isolation and characterization of phage-host systems from the Baltic Sea ice. Extremophiles. 18:121-130.
• Mönttinen HAM, Ravantti JJ, Stuart DI, and Poranen MM. 2014. Automated structural comparisons clarify the phylogeny of the right-hand-shape polymerases. Mol Biol Evol. 31:2741-52.
• Pietilä MK, Demina TA, Atanasova NS, Oksanen HM, and Bamford DH. 2014. Archaeal viruses and bacteriophages: comparisons and contrasts. Trends in Microbiol. 22:334-344.
• Sun X, Bamford DH, Poranen MM. 2014. Electrostatic interactions drive the self-assembly and the transcription activity of the Pseudomonas phage Ф6 procapsid. J Virol. 88:7112-7116.

2013

• Atanasova NS, Pietilä M, and Oksanen HM. 2013. Diverse antimicrobial interactions of halophilic archaea and bacteria extend over geographical distances and cross the domain barrier. MicrobiologyOpen. 2:811-825.
• Badia-Martinez D, Oksanen HM, Stuart, DI, and Abrescia NGA. 2013. Combined approaches to study virus structures. In: Structure and Physics of Viruses: An intergrated Textbook, Subcellular Biochemistry 68. Ed: Mateu MG. Springer. pp. 203-246.
• Hanhijärvi KJ, Žiedaitė G, Pietilä MK, Haeggström E, and Bamford DH. 2013. DNA ejection from an archaeal virus - a single-molecule approach. Biophys. J. 104:2264-2272.
• Peralta B, Gil-Carton D, Castaño-Díez D, Bertin A, Boulogne C, Oksanen HM, Bamford DH, and Abrescia NGA. 2013. Mechanism of membranous tunnelling nanotube formation in viral genome delivery. PLoS Biol. 11: e1001667.
• Pietilä MK, Atanasova NS, Oksanen HM, and Bamford DH. 2013. Modified coat protein forms the flexible spindle-shaped virion of haloarchaeal virus His1. Environ. Microbiol. 15:1674-1686.
• Pietilä MK, Laurinmäki P, Russell DA, Ko C-C, Jacobs-Sera D, Butcher SJ, Bamford DH, and Hendrix RW. 2013. Insights into head-tailed viruses infecting extremely halophilic archaea. J. Virol. 87:5057-5079.
• Pietilä MK, Laurinmäki P, Russell DA, Ko C-C, Jacobs-Sera D, Hendrix RW, Bamford DH, and Butcher SJ. 2013. Structure of the archaeal head-tailed virus HSTV-1 compleates the HK97-fold story. Proc. Natl. Acad. Sci. USA. 110:10604-10609.
• Ravantti JJ, Bamford DH, and Stuart DI.  2013. Automatic comparison and classification of protein structures. J. Struct. Biol. 183:47-56.
• Senčilo A, Jacobs-Sera D, Russell DA, Ko C-C, Bowman CA, Atanasova NS, Österlund E, Oksanen HM, Bamford DH, Hatfull GF, and Roine E, and Hendrix RW. 2013. Snapshot of haloarchaeal tailed virus genomes. RNA Biol. 10:33-46.
• Sun X, Pirttimaa MJ, Bamford DH, and Poranen MM. 2013. Rescue of maturation off-pathway products in the assembly of Pseudomonas phage φ6. J. Virol. 87:13279-13286.
• Vitale R, Roine E, Bamford, DH, and Corcelli A. 2013. Lipid fingerprints of intact viruses by MALDI-TOF mass spectrometry. Biochim Biophys Acta. 1831:872-879.

2012

• Aalto AP, Bitto D, Ravantti JJ, Bamford DH, Huiskonen JT, and Oksanen HM. 2012. A snapshot of virus evolution in hypersaline environments from the characterization of a membrane-containing Salisaeta icosahedral phage 1. Proc. Natl. Acad. Sci. USA. 109: 7079–7084.
• Jaakkola ST, Penttinen RK, Vilén ST, Jalasvuori M,  Rönnholm G, Bamford JKH, Bamford DH, and Oksanen HM. 2012. Closely related archaeal Haloarcula hispanica icosahedral viruses HHIV-2 and SH1 have nonhomologous genes encoding host recognition functions. J. Virol. 86:4734-4742.
• Kandiba L, Aitio O, Helin J, Guan Z, Permi P, Bamford DH, Eichler J, and Roine E. 2012. Diversity in prokaryotic glycosylation: an archaeal-derived N-linked glycan contains legionaminic acid. Mol. Microbiol. 84:578-593.
• Oksanen HM, Domanska A, and Bamford DH. 2012. Monolithic ion exchange chromatography methods for virus purification. Virology. 434:271-277.
• Pietilä M, Atanasova NS, Manole V, Liljeroos L, Butcher SJ, Oksanen HM, and Bamford DH. 2012. Virion architecture unifies globally distributed pleolipoviruses infecting halophilic archaea. J. Virol. 86:5067-5079.
• Senčilo A, Paulin L, Kellner S, Helm M, and Roine E. 2012. Related haloarchaeal pleomorphic viruses contain different genome types. Nucleic Acids Res. 40:5523-5534.
• Sun X, Bamford DH, and Poranen MM. 2012. Probing, by self-assembly, the number of potential binding sites for minor protein subunits in the procapsid of double-stranded RNA bacteriophage φ6. J. Virol. 86:12208–12216.

Here are some reading tips for technicues utilized for biological macromolecular complexes

Ultracentrifugation

Mohr, H. and Völkl, A. (2017). Ultracentrifugation. In eLS, John Wiley & Sons, Ltd (Ed.). doi:10.1002/9780470015902.a0002969.pub3

Chromatography

Lewis, A. C. (2015). Chromatographic Techniques. In eLS, John Wiley & Sons, Ltd (Ed.). doi:10.1002/9780470015902.a0002705.pub2

Sheehan, D. and O'Sullivan, S. (2001). Ion Exchange Chromatography. In eLS, (Ed.). doi:10.1038/npg.els.0002677

Mansoor, M. A. (2015). Liquid Chromatography. In eLS, John Wiley & Sons, Ltd (Ed.). doi/10.1002/9780470015902.a0002679.pub3