29. Zhang J, Zhang D, Sun L, Ji G, Huang X,
Niu T, Sun F (2019) VHUT-cryo-FIB, a
method to fabricate frozen-hydrated lamella
of tissue specimen for in situ cryo-electron
tomography. bioRxiv. https://doi.org/10.
1101/727149
30. Mahamid J, Schampers R, Persoon H, Hyman
AA, Baumeister W, Plitzko JM (2015) A
focused ion beam milling and lift-out approach
for site-specific preparation of frozen-hydrated
lamellas from multicellular organisms. J Struct
Biol 192(2):262–269
31. Stevie FA, Griffis DP, Russell PE (2005)
Focused ion beam gases for deposition and
enhanced etch. Springer US, Boston, MA, pp
53–72
32. Diebolder C, Faas F, Koster A, Koning R
(2015) Conical fourier shell correlation applied
to electron tomograms. J Struct Biol 190
(2):215–223
33. Strunk K, Wang K, Ke D, Gray J, Zhang P
(2012) Thinning of large mammalian cells for
cryo-TEM characterization by cryo-FIB milling. J Microsc 247(3):220–227
34. Medeiros JM, Bo ¨ck D, Weiss GL, Kooger R,
Wepf RA, Pilhofer M (2018) Robust workflow
and instrumentation for cryo-focused ion beam
milling of samples for electron cryotomography. Ultramicroscopy 190:1–11
35. Guo YS, Furrer JM, Kadilak AL, Hinestroza
HF, Gage DJ, Cho YK, Shor LM (2018) Bacterial extracellular polymeric substances amplify
water content variability at the pore scale.
Front Environ Sci 6:93
36. Arnold J, Mahamid J, Lucic V, de Marco A,
Fernandez JJ, Laugks T, Mayer T, Hyman AA,
Baumeister W, Plitzko JM (2016) Site-specific
cryo-focused ion beam sample preparation
guided by 3D correlative microscopy. Biophys
J 110(4):860–869
37. Nahmani M, Lanahan C, DeRosier D, Turrigiano GG (2017) High-numerical-aperture
cryogenic light microscopy for increased precision of superresolution reconstructions. Proc
Natl Acad Sci 114(15):3832–3836
38. Carlson DB, Evans JE (2011) Low-cost cryolight microscopy stage fabrication for correlated light/electron microscopy. JoVE 52:
e2909
39. Tuijtel MW, Koster AJ, Jakobs S, Faas FGA,
Sharp TH (2019) Correlative cryo superresolution light and electron microscopy on
mammalian cells using fluorescent proteins.
Sci Rep 9(1):1369
40. Schellenberger P, Kaufmann R, Siebert CA,
Hagen C, Wodrich H, Gru ¨newald K (2014)
High-precision correlative fluorescence and
electron cryo microscopy using two independent alignment markers. Ultramicroscopy
143:41–51. SI: Correlative microscopy
41. Chang YW, Chen S, Tocheva EI, TreunerLange A, Lo ¨bach S, Søgaard-Andersen L, Jensen GJ (2014) Correlated cryogenic photoactivated localization microscopy and cryoelectron tomography. Nat Methods 11
(7):737–739
42. Kaufmann R, Schellenberger P, Seiradake E,
Dobbie IM, Jones EY, Davis I, Hagen C, Gru ¨newald K (2014) Super-resolution microscopy
using standard fluorescent proteins in intact
cells under cryo-conditions. Nano Lett 14
(7):4171–4175
43. Wolff G, Limpens RWAL, Zheng S, Snijder EJ,
Agard DA, Koster AJ, Ba ´rcena M (2019) Mind
the gap: micro-expansion joints drastically
decrease the bending of FIB-milled cryo-lamellae. J Struct Biol 208:107389
44. Mastronarde DN (2005) Automated electron
microscope tomography using robust prediction of specimen movements. J Struct Biol
152(1):36–51
45. Schorb M, Haberbosch I, Hagen WJH,
Schwab Y, Mastronarde DN (2019) Software
tools for automated transmission electron
microscopy. Nat Methods 16(6):471–477
46. Marko M, Hsieh C, Moberlychan W, Mannella
CA, Frank J (2006) Focused ion beam milling
of vitreous water: prospects for an alternative to
cryo-ultramicrotomy
of
frozen-hydrated
biological samples. J Microsc 222(1):42–47
47. Toro-Nahuelpan M, Zagoriy I, Senger F,
Blanchoin L, The ´ry M, Mahamid J (2020) Tailoring cryo-electron microscopy grids by
photo-micropatterning for in-cell structural
studies. Nat Methods 17:50–54
48. Stokes DJ, Vystavel T, Morrissey F (2007)
Focused ion beam (FIB) milling of electrically
insulating specimens using simultaneous primary electron and ion beam irradiation. J
Phys D Appl Phys 40(3):874–877
82
Vinson Lam and Elizabeth Villa
Niu T, Sun F (2019) VHUT-cryo-FIB, a
method to fabricate frozen-hydrated lamella
of tissue specimen for in situ cryo-electron
tomography. bioRxiv. https://doi.org/10.
1101/727149
30. Mahamid J, Schampers R, Persoon H, Hyman
AA, Baumeister W, Plitzko JM (2015) A
focused ion beam milling and lift-out approach
for site-specific preparation of frozen-hydrated
lamellas from multicellular organisms. J Struct
Biol 192(2):262–269
31. Stevie FA, Griffis DP, Russell PE (2005)
Focused ion beam gases for deposition and
enhanced etch. Springer US, Boston, MA, pp
53–72
32. Diebolder C, Faas F, Koster A, Koning R
(2015) Conical fourier shell correlation applied
to electron tomograms. J Struct Biol 190
(2):215–223
33. Strunk K, Wang K, Ke D, Gray J, Zhang P
(2012) Thinning of large mammalian cells for
cryo-TEM characterization by cryo-FIB milling. J Microsc 247(3):220–227
34. Medeiros JM, Bo ¨ck D, Weiss GL, Kooger R,
Wepf RA, Pilhofer M (2018) Robust workflow
and instrumentation for cryo-focused ion beam
milling of samples for electron cryotomography. Ultramicroscopy 190:1–11
35. Guo YS, Furrer JM, Kadilak AL, Hinestroza
HF, Gage DJ, Cho YK, Shor LM (2018) Bacterial extracellular polymeric substances amplify
water content variability at the pore scale.
Front Environ Sci 6:93
36. Arnold J, Mahamid J, Lucic V, de Marco A,
Fernandez JJ, Laugks T, Mayer T, Hyman AA,
Baumeister W, Plitzko JM (2016) Site-specific
cryo-focused ion beam sample preparation
guided by 3D correlative microscopy. Biophys
J 110(4):860–869
37. Nahmani M, Lanahan C, DeRosier D, Turrigiano GG (2017) High-numerical-aperture
cryogenic light microscopy for increased precision of superresolution reconstructions. Proc
Natl Acad Sci 114(15):3832–3836
38. Carlson DB, Evans JE (2011) Low-cost cryolight microscopy stage fabrication for correlated light/electron microscopy. JoVE 52:
e2909
39. Tuijtel MW, Koster AJ, Jakobs S, Faas FGA,
Sharp TH (2019) Correlative cryo superresolution light and electron microscopy on
mammalian cells using fluorescent proteins.
Sci Rep 9(1):1369
40. Schellenberger P, Kaufmann R, Siebert CA,
Hagen C, Wodrich H, Gru ¨newald K (2014)
High-precision correlative fluorescence and
electron cryo microscopy using two independent alignment markers. Ultramicroscopy
143:41–51. SI: Correlative microscopy
41. Chang YW, Chen S, Tocheva EI, TreunerLange A, Lo ¨bach S, Søgaard-Andersen L, Jensen GJ (2014) Correlated cryogenic photoactivated localization microscopy and cryoelectron tomography. Nat Methods 11
(7):737–739
42. Kaufmann R, Schellenberger P, Seiradake E,
Dobbie IM, Jones EY, Davis I, Hagen C, Gru ¨newald K (2014) Super-resolution microscopy
using standard fluorescent proteins in intact
cells under cryo-conditions. Nano Lett 14
(7):4171–4175
43. Wolff G, Limpens RWAL, Zheng S, Snijder EJ,
Agard DA, Koster AJ, Ba ´rcena M (2019) Mind
the gap: micro-expansion joints drastically
decrease the bending of FIB-milled cryo-lamellae. J Struct Biol 208:107389
44. Mastronarde DN (2005) Automated electron
microscope tomography using robust prediction of specimen movements. J Struct Biol
152(1):36–51
45. Schorb M, Haberbosch I, Hagen WJH,
Schwab Y, Mastronarde DN (2019) Software
tools for automated transmission electron
microscopy. Nat Methods 16(6):471–477
46. Marko M, Hsieh C, Moberlychan W, Mannella
CA, Frank J (2006) Focused ion beam milling
of vitreous water: prospects for an alternative to
cryo-ultramicrotomy
of
frozen-hydrated
biological samples. J Microsc 222(1):42–47
47. Toro-Nahuelpan M, Zagoriy I, Senger F,
Blanchoin L, The ´ry M, Mahamid J (2020) Tailoring cryo-electron microscopy grids by
photo-micropatterning for in-cell structural
studies. Nat Methods 17:50–54
48. Stokes DJ, Vystavel T, Morrissey F (2007)
Focused ion beam (FIB) milling of electrically
insulating specimens using simultaneous primary electron and ion beam irradiation. J
Phys D Appl Phys 40(3):874–877
82
Vinson Lam and Elizabeth Villa