ISSN:
1615-6102
Keywords:
Laser scanning confocal microscopy
;
Multiple cell layers
;
Plant microtubules
;
Plant microfilaments
;
Roots
;
Tissue clearing
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
Notes:
Summary A protocol was developed to observe plant microtubules and actin microfilaments in large tissue samples without physical sectioning. Rye (Secale cereale L. cv. Rymin) root tip pieces from two-day-old seedlings were fixed and processed for immunolabeling. Incubation times of 24–48 h were required to insure adequate penetration of fixatives, antibodies, and washing buffers. Clearing of the tissue with methyl salicylate reduced background auto-fluorescence that would otherwise interfere with the resolution of cytoskeletal structures. Microtubules or microfilaments in 5–7 cell layers were visualized using the optical-sectioning capability of laser scanning confocal microscopy (LSCM) and projected as three-dimensional images. The three-dimensional character of the cytoskeletal elements is retained when viewing stained cells of intact tissue. The net-like character of a microfilament array radiating out from a single point into the cytoplasm is maintained when the cells are stained in intact root tip pieces and imaging is accomplished in situ.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF01281819
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