All of the equipment required for wholemount muscle dissection have been described previously (Sleigh et al., 2014a (link)). These or similar tools can also be used to prepare the ETA for in vivo imaging, while a description of how to induce anaesthesia and perform intramuscular injections has been detailed elsewhere (Sleigh et al., 2020b (link); Turney et al., 2012 (link)). Information on the primary and secondary antibodies used for immunofluorescence are provided in Tables 1 and 2 , respectively. AlexaFluor 488 and 555 α‐bungarotoxin (α‐BTX, Life Technologies, B13422 and B35451/RRID:AB_2617152, respectively, 1:1000) were used to identify postsynaptic acetylcholine receptors (AChRs). The binding fragment of tetanus neurotoxin (HCT) was produced and labelled with AlexaFluor 555 C2 maleimide (Life Technologies, A20346) as previously described (Gibbs et al., 2016 (link)). All dissection images and videos were taken using a DSK 500 dual head stereo microscope (Motic, Barcelona, Spain, PM5539B901) with attached Moticam 1080 HDMI digital camera (Motic, MC1080). Fixed and live immunofluorescent images were taken on an inverted LSM780 laser scanning microscope (Zeiss) using a 20×, 40× or 63× objective.
Dsk 500 dual head stereo microscope
Manufactured by Motic
Sourced in Spain
The DSK 500 is a dual head stereo microscope designed for laboratory use. It features two independent eyepiece tubes, allowing for simultaneous observation by two users. The microscope provides a wide range of magnification options to accommodate various sample types and observation requirements.
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3 protocols using dsk 500 dual head stereo microscope
1
Wholemount Muscle Dissection and Imaging
2
Dissection and Preparation of Murine Dorsal Root Ganglia
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The animals used in this protocol were 28 day-old female C57BL6/J mice, which were transcardially perfused with phosphate buffered saline (PBS) to decrease blood contamination (optional step). Mice can also be perfusion-xed if taking DRG for immuno uorescence analysis. The dissection can be performed from about post-natal day 5 (P5) onwards, with alternative options available for embryonic DRG [23, 24] . All images and videos (Additional Files 2-4) were taken using a DSK 500 dual head stereo microscope (Motic, Barcelona, Spain, PM5539B901) with attached Moticam 1080 HDMI digital camera (Motic, MC1080), except for Figure 1a,b,d, and e, and video 1 (Additional File 1), which were recorded using an iPhone 6s (Apple, Cupertino, CA). To preserve neuronal health for downstream applications, all steps should be completed e ciently and tissue kept as cold as possible. Use of a laminar ow hood will reduce contamination frequency of primary cultures [19] , but is not strictly necessary.
3
Dissection and Isolation of Murine DRG
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The animals used in this protocol were 28 day-old female C57BL6/J mice, which were transcardially perfused with phosphate buffered saline (PBS) to decrease blood contamination (optional step). Mice can also be perfusion-xed if taking DRG for immuno uorescence analysis. The dissection can be performed from about post-natal day 5 (P5) onwards, with alternative options available for embryonic DRG [23, 24] . All images and videos (Additional Files 2-4) were taken using a DSK 500 dual head stereo microscope (Motic, Barcelona, Spain, PM5539B901) with attached Moticam 1080 HDMI digital camera (Motic, MC1080), except for Fig. 1a,b,d, and e, and video 1 (Additional File 1), which were recorded using an iPhone 6 s. To preserve neuronal health for downstream applications, all steps should be completed e ciently and tissue kept as cold as possible. Use of a laminar ow hood will reduce contamination frequency of primary cultures [19] , but is not strictly necessary.
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