CryoEM

• Basic and advanced cryoEM techniques
• Project planning and assessment
• Sample optimisation and preparation
• Plunge freezing services (BSL2)
• Automated single particle and cryo-tomography digital data collection
• Advice on image reconstruction
• Collaborative projects
• Courses in structural biology, cryo electron microscopy and image reconstruction

Cryo-EM is applicable for projects in e.g. structural biology,nanotechnology,time-resolved assembly and phase transitions. Depending on the approach used, subnanometer resolution can be achieved.

• Single particle averaging and three dimensional (3D) reconstructions
• Electron tomography and 3D-reconstructions
• Measurements and statistical analyses
• Study of biological macromolecular complexes and other electron-beam sensitive materials in aqueous solutions.
• Particle sizes that range from ~10 to 300 nm in diameter.

Leica vitrification robot EM GP

• Automatic plunge freezer for the bare grid technique that is used for the preparation of vitrified fluid samples or extremely thin samples for cryo-EM.
• Suitable for biological suspensions and industrial emulsions in both, aqueous and inorganic solvents.
• Can be used to plunge freeze samples not only on EM grids, but also sapphire discs and samples in freeze fracture planchettes.
• Temperature and humidity controlled.
• Biosafety laboratory level 2

Manual vitrification guillotine in

• Biosafety laboratory level 2

Linkam-Zeiss correlative fluorescence light microscopy for vitrified specimens.

• For detection of fluorescence in vitrified specimens prior to EM. The region of interest can be rapidly located and then used for intance in electron tomography.

FEI TALOS Artica transmission electron microscope equipped with a Falcon III direct electron detector camera and FEI phase plate.

• FEI TALOS Arctica transmission electron microscope with a direct electron detector and a phase plate
• Improved signal-to-noise ratio, greater sensitivity and automated data collection (24/7) enabling high resolution cryo-EM.
• Only system available in Finland and is one of the few in Europe.
• Advanced techniques available include correlative light and electron microscopy for vitrified specimens (CLEM).
• Data can be transferred to the CSC Ltd for processing of frames, or returned on a USB drive.

• Cryo EM services are open to both academic and commercial users.
• Projects will undergo scientific review for their feasibility.

Instructions how to start a new project (9.1.2017):

1. Contact Pasi Laurinmäki or Benita Löflund (grp-cryoemservice at helsinki dot fi), or the Instruct-FI hub coordinator (grp-InstructHilifeHub at helsinki dot fi).
2. Have a kickoff meeting.
3. Fill in an application form for a new project. This will help scientific review of your project. If accepted, you will receive a unique ID that you will use for reservations, samples etc.  
4. Once sample delivery has been agreed, fill in the Sample sheet (link, see below) for sample delivery using your unique identifier. This will be used for billing, so keep the number to yourself.
5. Samples are brought to Biocenter 1, EM unit floor and delivered to Pasi Laurinmäki or Benita Löflund at an agreed date and time.
6. Samples are made and images are collected and delivered to the customer.

• Project sheet 2020 (application form)
• Sample sheet 2020

ACKNOWLEDGEMENT

Please notice that by using cryo EM service academic customers commit to acknowledge the unit in their publications or theses.  Please remember to add following sentences in your publication. We highly appreciate it.

"We thank Benita Löflund and Pasi Laurinmäki (University of Helsinki) for technical assistance in cryoEM. The facilities and expertise of the HiLIFE CryoEM unit at the University of Helsinki, a member of Instruct-ERIC Centre Finland, FINStruct, and Biocenter Finland are gratefully acknowledged."

CryoEM services offers full-service and self-service options for CryoEM work. Full-service option covers both sample preparation and imaging. Self-service customers will submit ready-made grids to our facility and imaging will be taken care of by our staff. Prices for both categories are given below.

All prices are VAT-exclusive. Standard rate of VAT is 24%. This price list is only for academic users. Prices for commercial customers are available on request.

Billing is three times a year.

FULL-SERVICE

Including Sample preparation and data collection as service.

Sample Preparation by Plunge freezing technique

• Standard grid 99,90 €
• Sample preparation service includes preparation of 3 parallel cryo-grids (Quantifoil Cu R1.2/1.3 grids and consumables) made by cryo-EM-staff. The grids will be stored for 6 months and they will be discarded without further notification.
• If other types of grids are requested and are available, the price will be obtained on request.
• Price includes: Preparation of EM-samples by plunge freezing technique, storage of cryo-sample for 6 months, consumables.

Negative staining

• This technique is used for sample optimization at room temperature. Cryo-EM-unit provides this method only as an additional service for optimisation of samples for cryo-EM. Price is available on request. The EM unit (Jokitalo) provides negative staining for non-cryo related projects as a regular service.

Data collection with Talos Arctica

Quick-check: 220 €/sample

• Used for sample optimization. Data set of the sample is typically 20 - 150 images. Such a data set will give you general information about your sample such as purity, size distribution and concentration.
• Price includes: Instrument time (Talos Arctica), operator and consumables. Sample preparation is not included.

Sample characterization: 440€/sample

• Representative data set of images. Data set of the sample is typically 100 - 300 images.
• Price includes: Instrument time (Talos Arctica), operator and consumables. Sample preparation is not included.

Large scale data collection for cryo-sample

• Large enough data set for particle reconstruction.

  • 24h, 1016 €. Data set of the sample is typically 500 - 1000 images.

  • 72h, 2744 €. Data set of the sample is typically 1000 - 3000 images.

• Price includes: Instrument time (Talos Arctica), operator and consumables. Sample preparation is not included.

Tomography (cryo)

• This technique
• Tilt series aquisition is

Instrument time

• Talos Arctica 8 am – 4 pm 55 € / h, 4 pm – 8 am 36 € / h
• Leica EM GP 8 € / shift
• Glow discharge 3,30 € / use

Grids

• Quantifoil R2/2 Au Finder 6,00 €
• Quantifoil R2/2 Au 200Mesh 7,70 €
• Ultrathin carbon film on holey carbon 5,20 €
• C-flat 8,10 €
• UltraAuFoil R2/2 7,50 €

Consumables

• Autogrid 12,70 €
• Autogrid Container 20,65 €
• Operator time 39,00 €

SELF-SERVICE

Customer submit cryo grids, no sample preparation service included.

Sample preparation

• Leica EM GP 8 € / h, minimum booking time is 2h. Price includes blotting paper, ethane and LN2.
• Glow discharge 3,30 € / use

Purchase of grids and grid containers is customer’s responsibility. If the grid containersupplied by the customer is not suitable for autogrids, grids will not be saved after data collection.

Storage and book keeping of cryogrids is the customer’s responsibility. Sample sheet needs to be filled in for all samples/grids that are stored in CryoEM facilities or submitted to CryoEM service for imaging.

DATA COLLECTION, Talos Arctica

Quick-check:                 

Used for sample optimization. Data set of the sample is typically 20 - 150 images. Such a data set will give you general information about your sample such as purity, size distribution and concentration.

• Instrument time 220 €/sample⁽¹
• Autogrid consumables 38 €/sample^{(1, 2, 3}

Sample includes max 3 identical replica grids (identical concentration, grid type, sample preparation parameters etc.) and data will be collected from first suitable grid. Other grids will not be checked.

Sample characterization:

Representative data set of images. Data set of the sample is typically 100 - 300 images.

• Instrument time 440 €/sample⁽¹
• Autogrid consumables 38 €/sample^{(1, 2, 3}

Sample includes max 3 identical replica grids and data will be collected from first suitable grid. Other grids will not be checked.

Large scale data collection for cryo-sample

Typically large enough data set for particle reconstruction.

• Instrument time 1016 € / 24h       Data set of the sample is typically 500 - 1000 images.
• Instrument time 2744 € / 72h       Data set of the sample is typically 1000 - 3000 images.
• Autogrid consumables 12,70 €/grid (If used)

Cryo electron tomography

In tomography techique the specimen is tilted in the microscope to produce image series. Each image of the series is from same location and the images provide different orientations of the particles.  

• Instrument time 1016 €
• Sample preparation not included
• Autogrid consumables 12,70 €/grid (If used)

MICROSCOPY CONSUMABLES

• Autogrid consumables 38 €/sample^{(1, 2, 3}
• Autogrid container 22€/sample⁽⁴

1. Sample includes max 3 identical replica grids (identical concentration, grid type, sample preparation conditions etc.)
2. If customer submits autogrids, autogrid fee will not be charged
3. If sample is loaded into the microscope, but it is not imaged, Autogrid fee will be charged.
4. We can provide autogrid containers only if we have extra containers in storage.

Useful links about cryoEM

•  The talks from NRAMM Workshop on Advanced Topics in EM Structure Determination: Challenges and Opportunitiesare available on-line at: http://nramm.nysbc.org/2017-workshop-lectures/. Topics discussed  included specimen preparation, imaging, instrumentation, processing and reconstruction, and methods for validation. 

Recent publications

2021

• Flatt, J.W.,  Domanska, A., Seppälä, A.L., Butcher, S.J. (2021) Identification of a conserved virion-stabilizing network inside the interprotomer pocket of enteroviruses. Commun Biol 4:250  https://doi.org/10.1038/s42003-021-01779-x

2020

• Barreiro, K., Dwivedi, O. P., Leparc, G., Rolser, M., Delic, D., Forsblom, C., Groop, P. H., Groop, L., Huber, T. B., Puhka, M., & Holthofer, H. (2020). Comparison of urinary extracellular vesicle isolation methods for transcriptomic biomarker research in diabetic kidney disease. Journal of extracellular vesicles, 10(2), e12038. https://doi.org/10.1002/jev2.12038
• Mamata Bhattarai, Fabio Valoppi, Sami-Pekka Hirvonen, Sami Hietala, Petri Kilpeläinen, Vladimir Aseyev, Kirsi S. Mikkonen (2020) Time-dependent self-association of spruce galactoglucomannans depends on pH and mechanical shearing. Food Hydrocolloids Volume 102. https://doi.org/10.1016/j.foodhyd.2019.105607.
• Heikki Saari,Tiia Turunen,Andres Lõhmus,Mikko Turunen,Matti Jalasvuori,Sarah J. Butcher,Seppo Ylä-Herttuala,Tapani Viitala,Vincenzo Cerullo,Pia R. M. Siljander &Marjo Yliperttula. (2020) Extracellular vesicles provide a capsid-free vector for oncolytic adenoviral DNA delivery.Journal of Extracellular Vesicles, 9:1, 1747206, https://doi.org/10.1080/20013078.2020.1747206

2019

• Abdelnabi, R., Geraets, J.A., Ma, Y, Mirabelli, C., Flatt, J.W. , Domanska, A., Delang, L., Jochmans, D., Jayaprakash, V.,  Sinha, B.N.,  Leyssen, P., Butcher, S.J., Neyts , J. (2019) A novel druggable interprotomer pocket in the capsid of rhino- and enteroviruses. PLoS Biology 17: e3000281. https://doi.org/10.1371/journal.pbio.3000281
• Domanska, A. Flatt, J.W., Jukonen, J.J.J., Geraets, J.A., Butcher, S.J.  (2019) 2.8 Å resolution cryo-EM structure of human parechovirus 3 in complex with Fab from a neutralizing antibody. J. Virol   93:e01597-18. doi: 10.1128/JVI.01597-18
• Pooch F, Sliepen M, Knudsen KD, Nyström B, Tenhu H, Winnik FM. (2019) Poly(2-isopropyl-2-oxazoline)-b-poly(lactide) (PiPOx-b-PLA) Nanoparticles in Water: Interblock van der Waals Attraction Opposes Amphiphilic Phase Separation. Macromolecules. 2019 Feb 12;52(3):1317-1326. doi: 10.1021/acs.macromol.8b02558. Epub 2019 Feb 1. PMID: 31496543
• Ruokolainen V., Domanska, A., Pellicia, M., Laajala, M., Butcher, S.J., Marjomäki, M. (2019) Extracellular albumin and endosomal ions prime enterovirus particles for uncoating that can be prevented by fatty acid saturation. J. Virol. 93:e01597-18  doi: 10.1128/JVI.01597-18 https://doi.org/10.1128/JVI.00599-19

• Sah-Teli SK, Hynönen MJ, Schmitz W, Geraets JA, Seitsonen J, Pedersen JS, Butcher SJ, Wierenga RK, Venkatesan R. (2019) Complementary substrate specificity and distinct quaternary assembly of the Escherichia coli aerobic and anaerobic β-oxidation trifunctional enzyme complexes. Biochem J. 2019 Jul 15;476(13):1975-1994. doi: 10.1042/BCJ20190314. PMID: 31235482

2018

• Yang, D., Viitasuo, M., Pooch, F., Tenhu, H., Hietala, S. (2018) Poly(N-acryloylglycinamide) microgels as nanocatalyst platform. Polymer Chemistry. 9:4 517-524 doi: 10.1039/c7py01950e
• Hepojoki J, Hepojoki S, Smura T, Szirovicza L, Dervas E, Prähauser B, Nufer L, Schraner EM, Vapalahti O, Kipar A, Hetzel U. (2018). Characterization of Haartman Institute snake virus-1 (HISV-1) and HISV-like viruses-The representatives of genus hartmanivirus, family Arenaviridae. PLoS Pathog. 14:e1007415.

2017

• Russo G, Witos J, Rantamäki AH, Wiedmer SK. (2017) Cholesterol affects the interaction between an ionic liquid and phospholipid vesicles. A study by differential scanning calorimetry and nanoplasmonic sensing. Biochim Biophys Acta. 2017 Dec;1859(12):2361-2372. doi: 10.1016/j.bbamem.2017.09.011. Epub 2017 Sep 11.

2016

• Leon-Velarde CG, Happonen L, Pajunen M, Leskinen K, Kropinski AM, Mattinen L, Rajtor M, Zur J, Smith D, Chen S, Nawaz A, Johnson RP, Odumeru JA, Griffiths MW, Skurnik M. Yersinia enterocolitica-specific infection by bacteriophages TG1 and ϕR1-RT is dependent on temperature-regulated expression of the phage host receptor OmpF. Appl Environ Microbiol. 2016 Aug 15;82(17):5340-53. doi: 10.1128/AEM.01594-16
• Shakeel S., Westerhuis B.M., Domanska, A., Konig, R.I., Matadeen, R., Koster, A.J., Bakker, A.Q., Beaumont, T., Wolthers, K.C., Butcher, S.J. (2016) Multiple capsid-stabilizing interactions revealed in a high-resolution structure of an emerging picornavirus causing neonatal sepsis. Nature Communications 7:11387. doi: 10.1038/ncomms11387.

2015

• Guryanov, S., Liljeroos, L. Kasaragod, P., Kajander, T., Butcher, S.J. (2015) Crystal structure of the measles virus nucleoprotein core in complex with an N-terminal region of phosphoprotein. J. Virol. 90:2849-57. doi: 10.1128/JVI.02865-15
• Shakeel S., Westerhuis B.M., Ora, A., Koen, G., Bakker, A.Q., Claassen, Y., Wagner, K., Beaumont, T., Wolthers, K.C., Butcher, S.J. (2015) Structural basis of human parechovirus neutralization by human monoclonal antibodies. J. Virol. 89:9571-80

2014

• Magarkar A, Mele N, Abdel-Rahman N, Butcher S, Torkkeli M, Serimaa R, Paananen A, Linder M, Bunker A. (2014) Hydrophobin film structure for HFBI and HFBII and mechanism for accelerated film formation. PLOS Computational Biology 10.1371/journal.pcbi.1003745
• Vahokoski J, Bhargav SP, Desfosses A, Andreadaki M, Kumpula EP, Martinez SM, Ignatev A, Lepper S, Frischknecht F, Sidén-Kiamos I, Sachse C, Kursula I. Structural differences explain diverse functions of Plasmodium actins. PLoS Pathog. 2014 Apr 17;10(4):e1004091.

2013

• Hedegaard, S., Nilsson, C., Laurinmäki, P., Butcher, S.J., Urtti, A., Yaghmur, A. (2013) Nanostructured aqueous dispersions of citrem interacting with lipids and PEGylated lipids. RSC Advances 3:24576–24585.
• Hetzel,U., Sironen,T., Laurinmäki, P., Liljeroos, L., Patjas, A., Henttonen, H., Vaheri, A., Artelt, A., Kipar, A., Butcher, S.J., Vapalahti O., Hepojoki J. (2013) Isolation, identification and characterization of novel Arenaviruses, the etiological agent of Boid Inclusion Body Disease. J. Virol. 87:20 10918-10935
• Hirvonen, S, Karesoja, M., Karjalainen, E., Hietala, S., Laurinmäki, P., Vesanen, E., Butcher, S.J., Tenhu, H. (2013) Colloidal properties and gelation of aqueous dispersions of conductive poly(benzimidazobenzophenanthroline) derivatives. Polymer 54:694-701
• Karjalainen, E., Chenna, N., Laurinmäki, P., Butcher, S.J., Tenhu H. (2013) Diblock copolymers consisting of polymerized ionic liquid and poly(N-isopropylacrylamide). Effects of PNIPAM block length and counter ion on self-assembling and thermal properties.  Polym.Chem. 4:1014-1024
• Nilsson, C., Barrios-Lopez, B., Kallinen, A., Laurinmӓki, P., Butcher, S.J., Raki, M., Bergstrӧm, K., Weng Larsen, S., Østergaard, J., Larsen, C., Urtti, A., Airaksinen, A., Yaghmur, A. (2013) SPECT/CT imaging of radiolabeled cubosomes and hexosomes for potential theranostic applications. Biomaterials 34:8491-8503
• Pietilä, M.K., Laurinmäki, P., Russell, D.A., Ko, C., Jacobs-Sera, D., Butcher, S.J., Bamford, D.H., Hendrix, R.W. (2013) Insights into head-tailed viruses infecting extremely halophilic archaea. J. Virol. 87:3248-3260
• Pietilä, M.K., Laurinmäki, P., Russell, D.A., Ko, C., Jacobs-Sera, D., Hendrix, R.W., Bamford, D.H., Butcher, S.J.. (2013) Structure of the archaeal head-tailed virus HSTV-1 completes the HK97-fold story. Proc. Natl. Acad. Sci. (USA) 110:10604-10609

2012

• Dearborn, A.D., Laurinmäki, P., Chandramouli, P., Rodenburg, C.M., Wang, S., Butcher, S.J., Dokland, T. (2012) Structure and size determination of bacteriophage P2 and P4 procapsids: function of size responsiveness mutations. J. Struct. Biol. 178:215-224
• Koho, T., Mäntylä,T., Laurinmäki, P., Huhti, L. Butcher, S., Vesikari,T., Kulomaa, M.S., Hytönen, V.P. (2012) Purification of norovirus-like particles (VLPs) by ion exchange chromatography. J. Vir. Methods. 181:6-11.
• Pietilä, MK, Atanasova, NS, Manole, V, Liljeroos, L., Butcher, SJ, Oksanen, HM, Bamford, DH. (2012) Virion architecture unifies globally distributed pleolipoviruses infecting halophilic archaea. J. Virol, 86:5067-5079
• Sarin, L.P, Hirvonen, J., Laurinmäki, P., Butcher, S.J., Bamford, D.H., Poranen, M.M. (2012) Bacteriophage ϕ6 nucleocapsid surface protein 8 interacts with virus-specific membrane vesicles containing the major envelope protein 9. J. Virol. 86:5376-5379
• M. Sedlák: Homopolymer Self-assembly into Stable Nanoparticles: Concerted Action of Hydrophobic Association and Hydrogen Bonding in Thermoresponsive Poly(alkylacrylic acid)s, J. Phys. Chem. B, 116 (8), 2356–2364, 2012.
• Skurnik, M., Hyytiäinen, H., Happonen, L., Kiljuenn, S., Datta, N., Mattinen, L., Williamson, K., Kristo, P., Szeliga, M., Kalin-Mänttätri, L., Ahola-Iivarinen, E., Kalkkinen, N., Butcher S.J. (2012) Characterization of the genome, proteome and structure of yersiniophage φR1-37. J. Virol. 86:12625-12642

2011

• Alhoranta, A., Lehtinen, J., Urtti, A., Butcher, S.J., Aseyev, V., Tenhu, H. (2011) Cationic amphiphilic star and linear block copolymers: synthesis, self-assembly and in vitro gene transfection. Biomacromolecules 12;3213
• Koho, T., Huhti, L., Blazevic, V., Nurminen, K., Butcher, S.J., Laurinmäki, P., Kalkkinen, N., Rönnholm, G., Vesikari, T., Hytönen, V.P., Kulomaa, M.S. (2011) Production and characterization of virus-like particles and the P domain protein of GII.4 norovirus. J. Vir. Methods doi:10.1016/j.jviromet.2011.05.009
• Kumar, V., Butcher, S.J., Öörni, K., Engelhardt, P., Heikkonen, J., Kaski, K., Ala-Korpela, M., Kovanen, P.T.(2011) Three-dimensional cryoEM reconstruction of native LDL particles to 16å resolution at physiological body temperature PLoS ONE 6(5): e18841. doi:10.1371/journal.pone.0018841.