Materials List for
Whole-Mount Immunofluorescence Staining, Confocal
Imaging and 3D Reconstruction of the Sinoatrial and
Atrioventricular Node in the Mouse
Ruibing Xia1,2,3, Julia Vlcek1,2, Julia Bauer1,2,3, Stefan Kääb1,3, Hellen Ishikawa-Ankerhold1,2, Dominic Adam van den
Heuvel1,2, Christian Schulz1,2,3, Steffen Massberg1,2,3, Sebastian Clauss1,2,3
1 2
University Hospital Munich, Department of Medicine I, Ludwig Maximilians University (LMU) Munich Institute of Surgical Research at the Walter Brendel
3
Center of Experimental Medicine, University Hospital Munich, Ludwig Maximilians University (LMU) Munich German Center for Cardiovascular Research
(DZHK), Partner Site Munich, Munich Heart Alliance
Corresponding Author Citation
Ruibing Xia Xia, R., Vlcek, J., Bauer, J., Kääb, S., Ishikawa-Ankerhold, H., van den Heuvel, D.A.,
[email protected] Schulz, C., Massberg, S., Clauss, S. Whole-Mount Immunofluorescence Staining, Confocal
Imaging and 3D Reconstruction of the Sinoatrial and Atrioventricular Node in the Mouse. J.
Vis. Exp. (166), e62058, doi:10.3791/62058 (2020).
Date Published DOI URL
December 22, 2020 10.3791/62058 jove.com/video/62058
Materials
Name Company Catalog Number Comments
Anesthesia
Isoflurane vaporizer system Hugo Sachs Elektronik 34-0458, 34-1030, 73-4911, 34-0415, Includes an induction chamber, a gas
73-4910 evacuation unit and charcoal filters
Modified Bain circuit Hugo Sachs Elektronik 73-4860 Includes an anesthesia mask for
mice
Surgical Platform Kent Scientific SURGI-M
In vivo instrumentation
Fine forceps Fine Science Tools 11295-51
Iris scissors Fine Science Tools 14084-08
Spring scissors Fine Science Tools 91500-09
Tissue forceps Fine Science Tools 11051-10
Tissue pins Fine Science Tools 26007-01 Could use 27G needles as a
substitute
General lab instruments
Orbital shaker Sunlab D-8040
Magnetic stirrer IKA RH basic
Pipette,volume 10 µL, 100 µL, 1000 Eppendorf Z683884-1EA
µL
Microscopes
Dissection stereo- zoom microscope VWR 10836-004
Laser Scanning Confocal microscope Zeiss LSM 800
Software
Imaris 8.4.2 Oxford instruments
ZEN 2.3 SP1 black Zeiss
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General Lab Material
0.2 µm syringe filter Sartorius 17597
100 mm petri dish Falcon 351029
27G needle BD Microlance 3 300635
50 ml Polypropylene conical Tube Falcon 352070
5ml Syringe Braun 4606108V
Cover slips Thermo Scientific 7632160
Eppendorf Tubes Eppendorf 30121872
Chemicals
0.5 M EDTA Sigma 20-158 Components of TEA
16% Formaldehyde Solution Thermo Scientific 28908 use as a 4% solution
Acetic acid Merck 100063 Components of TEA
Agarose Biozym 850070
Bovine Serum Albumin Sigma A2153-100G
DPBS (1X) Dulbecco's Phosphate Gibco 14190-094
Buffered Saline
Normal goat serum Sigma NS02L
Sucrose Sigma S1888-1kg
Tris-base Roche TRIS-RO Components of TEA
Triton X-100 Sigma T8787-250ml Diluted to 1% in PBS
Tween 20 Sigma P2287-500ml
Drugs
Fentanyl 0.5 mg/10 mL Braun Melsungen
Isoflurane 1 mL/mL Cp-pharma 31303
Oxygen 5 L Linde 2020175 Includes a pressure regulator
Antibodies
Goat anti-Rabbit IgG Alexa Cell Signaling Technology #4412 diluted to 1:200
Fluor 488
Goat anti-Rat IgG Alexa Fluor 647 Invitrogen #A-21247 diluted to 1:200
Hoechst 33342, Trihydrochloride, Invitrogen H3570 diluted to 1:1000
Trihydrate (DAPI)
Rabbit Anti-Connexin-43 Sigma C6219 diluted to 1:200
Rat anti-HCN4 (SHG 1E5) Invitrogen MA3-903 diluted to 1:200
Other
Plexiglass ring Self-designed and 3D printed
Plasticine Cernit 49655005
Silikonpasten, Baysilone VWR 291-1220
Animals
Mouse, C57BL/6 The Jackson Laboratory
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